3,4-dihydro-(1H)-quinazolin-2-ones and their use as CSBP/p38 kinase inhibitors

ABSTRACT

The present invention relates to 4,4-disubstituted-3,4-dihydro-2(1H)-quinazolines of formula (1), or stereoisomeric forms, stereoisomeric mixtures, or pharmaceutically acceptable salt forms thereof, which are useful as inhibitors of HIV reverse transcriptase, and to pharmaceutical compositions and diagnostic kits comprising the same, and methods of using the same for treating viral infection or as an assay standard or reagent.

This application is the §371 national stage entry of PCT/US/00/31908,filed Nov. 21, 2000, which claims benefit from provisional applicationU.S. Ser. No. 60/167,113 filed Nov. 23, 1999.

FIELD OF THE INVENTION

This invention relates to a novel group of 5-aryl-3,4-dihydro-(1H)quinazolin-2-one compounds, processes for the preparation thereof, theuse thereof in treating CSBP/p38 kinase mediated diseases andpharmaceutical compositions for use in such therapy.

BACKGROUND OF THE INVENTION

Intracellular signal transduction is the means by which cells respond toextracellular stimuli. Regardless of the nature of the cell surfacereceptor (e. g. protein tyrosine kinase or seven-transmembrane G-proteincoupled), protein kinases and phosphatases along with phopholipases arethe essential machinery by which the signal is further transmittedwithin the cell [Marshall, J. C. Cell, 80, 179-278 (1995)]. Proteinkinases can be categorized into five classes with the two major classesbeing, tyrosine kinases and serine/threonine kinases depending uponwhether the enzyme phosphorylates its substrate(s) on specifictyrosine(s) or serine/threonine(s) residues [Hunter, T., Methods inEnzymology (Protein Kinase Classification) p. 3, Hunter, T.; Sefton, B.M.; eds. vol. 200, Academic Press; San Diego, 1991].

For most biological responses, multiple intracellular kinases areinvolved and an individual kinase can be involved in more than onesignaling event. These kinases are often cytosolic and can translocateto the nucleus or the ribosomes where they can affect transcriptionaland translational events, respectively. The involvement of kinases intranscriptional control is presently much better understood than theireffect on translation as illustrated by the studies on growth factorinduced signal transduction involving MAP/ERK kinase [Marshall, C. J.Cell, 80, 179 (1995); Herskowitz, I. Cell, 80, 187 (1995); Hunter, T.Cell, 80, 225 (1995);Seger, R., and Krebs, E. G. FASEB J., 726-735(1995)].

While many signaling pathways are part of cell homeostasis, numerouscytokines (e.g., IL-1 and TNF) and certain other mediators ofinflammation (e.g., COX-2, and iNOS) are produced only as a response tostress signals such as bacterial lipopolysaccharide (LPS). The firstindications suggesting that the signal transduction pathway leading toLPS-induced cytokine biosynthesis involved protein kinases came fromstudies of Weinstein [Weinstein, et al., J. Immunol. 151, 3829(1993)]but the specific protein kinases involved were not identified. Workingfrom a similar perspective, Han [Han, et al., Science 265, 808(1994)]identified murine p38 as a kinase which is tyrosine phosphorylated inresponse to LPS. Definitive proof of the involvement of the p38 kinasein LPS-stimulated signal transduction pathway leading to the initiationof proinflammatory cytokine biosynthesis was provided by the independentdiscovery of p38 kinase by Lee [Lee; et al., Nature, 372, 739(1994)] asthe molecular target for a novel class of anti-inflammatory agents. Thediscovery of p38 (termed by Lee as CSBP 1 and 2) provided a mechanism ofaction of a class of anti-inflammatory compounds for which SK&F 86002was the prototypic example. These compounds inhibited IL-1 and TNFsynthesis in human monocytes at concentrations in the low uM range [Lee,et al., Int. J. Immunopharmac. 10(7), 835(1988)] and exhibited activityin animal models which are refractory to cyclooxygenase inhibitors [Lee;et al., Annals N. Y. Acad. Sci., 696, 149(1993)].

It is now firmly established that CSBP/p38 is a one of several kinasesinvolved in a stress-response signal transduction pathway which isparallel to and largely independent of the analogous mitogen-activatedprotein kinase (MAP) kinase cascade (FIG. 1). Stress signals, includingLPS, pro-inflammatory cytokines, oxidants, UV light and osmotic stress,activate kinases upstream from CSBP/p38 which in turn phosphorylateCSBP/p38 at threonine 180 and tyrosine 182 resulting in CSBP/p38activation. MAPKAP kinase-2 and MAPKAP kinase-3 have been identified asdownstream substrates of CSBP/p38 which in turn phosphorylate heat shockprotein Hsp 27 (FIG. 1). It is not yet known whether MAPKAP-2, MAPKAP-3,Mnk1 or Mnk2 are involved in cytokine biosynthesis or alternatively thatinhibitors of CSBP/p38 kinase might regulate cytokine biosynthesis byblocking a yet unidentified substrate downstream from CSBP/p38 [Cohen,P. Trends Cell Biol., 353-361(1997)].

What is known, however, is that in addition to inhibiting IL-1 and TNF,CSBP/p38 kinase inhibitors (SK&F 86002 and SB 203580) also decrease thesynthesis of a wide variety of pro-inflammatory proteins including,IL-6, IL-8, GM-CSF and COX-2. Inhibitors of CSBP/p38 kinase have alsobeen shown to suppress the TNF-induced expression of VCAM-1 onendothelial cells, the TNF-induced phosphorylation and activation ofcytosolic PLA2 and the IL-1-stimulated synthesis of collagenase andstromelysin. These and additional data demonstrate that CSBP/p38 isinvolved not only cytokine synthesis, but also in cytokine signaling[CSBP/P38 kinase reviewed in Cohen, P. Trends Cell Biol.,353-361(1997)].

Interleukin-1 (IL-1) and Tumor Necrosis Factor (TNF) are biologicalsubstances produced by a variety of cells, such as monocytes ormacrophages. IL-1 has been demonstrated to mediate a variety ofbiological activities thought to be important in immunoregulation andother physiological conditions such as inflammation [See, e.g.,Dinarello et al., Rev. Infect. Disease, 6, 51 (1984)]. The myriad ofknown biological activities of IL-1 include the activation of T helpercells, induction of fever, stimulation of prostaglandin or collagenaseproduction, neutrophil chemotaxis, induction of acute phase proteins andthe suppression of plasma iron levels.

There are many disease states in which excessive or unregulated IL-1production is implicated in exacerbating and/or causing the disease.These include rheumatoid arthritis, osteoarthritis, endotoxemia and/ortoxic shock syndrome, other acute or chronic inflammatory disease statessuch as the inflammatory reaction induced by endotoxin or inflammatorybowel disease; tuberculosis, atherosclerosis, muscle degeneration,cachexia, psoriatic arthritis, Reiter's syndrome, rheumatoid arthritis,gout, traumatic arthritis, rubella arthritis, and acute synovitis.Recent evidence also links IL-1 activity to diabetes and pancreatic βcells [review of the biological activities which have been attributed toIL-1 Dinarello, J. Clinical Immunology, 5 (5), 287-297 (1985)].

Excessive or unregulated TNF production has been implicated in mediatingor exacerbating a number of diseases including rheumatoid arthritis,rheumatoid spondylitis, osteoarthritis, gouty arthritis and otherarthritic conditions; sepsis, septic shock, endotoxic shock, gramnegative sepsis, toxic shock syndrome, adult respiratory distresssyndrome, cerebral malaria, chronic pulmonary inflammatory disease,silicosis, pulmonary sarcoisosis, bone resorption diseases, reperfusioninjury, graft vs. host reaction, allograft rejections, fever andmyalgias due to infection, such as influenza, cachexia secondary toinfection or malignancy, cachexia, secondary to acquired immunedeficiency syndrome (AIDS), AIDS, ARC (AIDS related complex), keloidformation, scar tissue formation, Crohn's disease, ulcerative colitis,or pyresis.

Interleukin-8 (IL-8) is a chemotactic factor produced by several celltypes including mononuclear cells, fibroblasts, endothelial cells, andkeratinocytes. Its production from endothelial cells is induced by IL-1,TNF, or lipopolysachharide (LPS). IL-8 stimulates a number of functionsin vitro. It has been shown to have chemoattractant properties forneutrophils, T-lymphocytes, and basophils. In addition it induceshistamine release from basophils from both normal and atopic individualsas well as lysozomal enzyme release and respiratory burst fromneutrophils. IL-8 has also been shown to increase the surface expressionof Mac-1 (CD11b/CD18) on neutrophils without de novo protein synthesis,this may contribute to increased adhesion of the neutrophils to vascularendothelial cells. Many diseases are characterized by massive neutrophilinfiltration. Conditions associated with an increased in IL-8 production(which is responsible for chemotaxis of neutrophil into the inflammatorysite) would benefit by compounds that are suppressive of IL-8production.

IL-1 and TNF affect a wide variety of cells and tissues and thesecytokines as well as other leukocyte derived cytokines are important andcritical inflammatory mediators of a wide variety of disease states andconditions. The inhibition of these cytokines is of benefit incontrolling, reducing and alleviating many of these disease states.

Inhibition of signal transduction via CSBP/p38, which in addition toIL-1, TNF and IL-8 described above is also required for the synthesisand/or action of several additional pro-inflammatory proteins (i.e.,IL-6, GM-CSF, COX-2, collagenase and stromelysin), is expected to be ahighly effective mechanism for regulating the excessive and destructiveactivation of the immune system. This expectation is supported by thepotent and diverse anti-inflammatory activities described for CSBP/p38kinase inhibitors [Badger, et al., J. Pharm. Exp. Thera 279 (3):1453-1461 (1996); Griswold, et al, Pharmacol. Comm. 7,323-229 (1996)].

There remains a need for treatment, in this field, for compounds whichare cytokine suppressive anti-inflammatory drugs, i.e. compounds whichare capable of inhibiting the CSBP/p38/RK kinase.

BRIEF DESCRIPTION OF THE DRAWINGS

The patent application filed contains at least one drawing executed incolor. Copies of this patent or patent application publication withcolor drawing(s) will be provided by the Patent & Trademark Office uponrequest and payment of necessary fee.

FIG. 1 demonstrates the p38 kinase cascade.

FIG. 2 demonstrates a Citric Acid Induced Cough Model.

FIG. 3 demonstrates an Antigen- or LTD4-Induced Hypertussive Model inthe Guinea Pig

FIG. 4 demonstrates Effects of Dextromethorphan or Codeine On CitricAcid-Induced Cough in Guinea Pigs.

SUMMARY OF THE INVENTION

This invention relates to the novel compounds of Formula (I), andpharmaceutical compositions comprising a compound of Formula (I), and apharmaceutically acceptable diluent or carrier.

This invention relates to a method of treating a CSBP/RK/p38 kinasemediated disease in a mammal in need thereof, which comprisesadministering to said mammal an effective amount of a compound ofFormula (I).

This invention also relates to a method of inhibiting cytokines and thetreatment of a cytokine mediated disease, in a mammal in need thereof,which comprises administering to said mammal an effective amount of acompound of Formula (I).

This invention more specifically relates to a method of inhibiting theproduction of IL-1 in a mammal in need thereof which comprisesadministering to said mammal an effective amount of a compound ofFormula (I).

This invention more specifically relates to a method of inhibiting theproduction of IL-6 in a mammal in need thereof which comprisesadministering to said mammal an effective amount of a compound ofFormula (I).

This invention more specifically relates to a method of inhibiting theproduction of IL-8 in a mammal in need thereof which comprisesadministering to said mammal an effective amount of a compound ofFormula (I).

This invention more specifically relates to a method of inhibiting theproduction of TNF in a mammal in need thereof which comprisesadministering to said mammal an effective amount of a compound ofFormula (I).

Accordingly, the present invention provides a compound of Formula (I):

wherein

R₁ is phenyl, naphth-1-yl, naphth-2-yl, heterocyclic or heteroaryl ring,which ring is optionally substituted independently by one to threesubstituents selected from halogen, C₁₋₄ alkyl, halo-substituted-C₁₋₂₄alkyl, cyano, nitro, (CR₁₀R₂₀)_(v)NR₄R₁₄, (CR₁₀R₂₀)_(v)C(Z)NR₄R₁₄,(CR₁₀R₂₀)_(v)C(Z)OR₈, (CR₁₀R₂₀)_(v)COR₃, (CR₁₀R₂₀)_(v)C(O)H, SR₅,S(O)R₅, S(O)₂R₅, (CR₁₀R₂₀)_(v)OR₈, ZC(Z)R₁₁, NR₁₀C(Z)R₁₁, orNR₁₀S(O)₂R₇;

R₂ is a C₁₋₁₀ alkyl, C₃₋₇ cycloalkyl, C₃₋₇cycloalkylC₁₋₁₀ alkyl, C₅₋₇cycloalkenyl, C₅₋₇ cycloalkenyl C₁₋₁₀ alkyl, aryl, arylC₁₋₁₀ alkyl,heteroaryl, heteroarylC₁₋₁₀ alkyl, heterocyclic, or heterocyclylC₁₋₁₀alkyl moiety, which moieties are optionally substituted one or moretimes independently with C₁₋₁₀ alkyl, halo-substituted C₁₋₁₀ alkyl,C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, C₃₋₇ cycloalkyl, C₃₋₇cycloalkylC₁₋₁₀alkyl, C₅₋₇ cycloalkenyl, C₅₋₇ cycloalkenyl C₁₋₁₀ alkyl, halogen,(CR₁₀R₂₀)_(n)OR₆, (CR₁₀R₂₀)_(n)SH, (CR₁₀R₂₀)_(n)S(O)_(m)R₇,(CR₁₀R₂₀)_(n)NHS(O)₂R₇, (CR₁₀R₂₀)_(n)NR₄R₁₄, (CR₁₀R₂₀)_(n)CN,(CR₁₀R₂₀)_(n)S(O)₂NR₄R₁₄, (CR₁₀R₂₀)_(n)C(Z)R₆, (CR₁₀R₂₀)_(n)OC(Z)R₆,(CR₁₀R₂₀)_(n)C(Z)OR₆, (CR₁₀R₂₀)_(n)C(Z)NR₄R₁₄, (CR₁₀R₂₀)_(n)NR₁₀C(Z)R₆,(CR₁₀C₂₀)_(n)NR₁₀C(═NR₁₀) NR₄R₁₄, (CR₁₀R₂₀)_(n)OC(Z)NR₄R₁₄,(CR₁₀R₂₀)_(n)NR₁₀C(Z) NR₄R₁₄, or (CR₁₀R₂₀)_(n)NR₁₀C(Z)OR₇;

R₃ is C₁₋₄ alkyl, halo-substituted C₁₋₄ alkyl, C₁₋₄ alkenyl, C₂₋₄alkynyl, C₃₋₇ cycloalkyl, C₅₋₇ cycloalkenyl, aryl, arylC₁₋₄ alkyl,heteroaryl, heteroarylC₁₋₄ alkyl, heterocyclyl, heterocyclylC₁₋₄ alkyl,(CR₁₀R₂₀)_(v)OR₇, (CR₁₀R₂₀)_(v)S(O)_(m)R₇, (CR₁₀R₂₀)_(v)NHS(O)₂R₇, or(CR₁₀R₂₀)_(v)NR₄R₁₄; and wherein the aryl, arylalkyl, heteroaryl,heteroaryl alkyl may be optionally substituted;

R₄ and R₁₄ is each independently selected from hydrogen or optionallysubstituted C₁₋₄ alkyl, optionally substituted aryl or optionallysubstituted aryl-C₁₋₄ alkyl, or together with the nitrogen which theyare attached form a heterocyclic ring of 5 to 7 members which ringoptionally contains an additional heteroatom selected from oxygen,sulfur or NR₉;

R₅ is hydrogen, C₁₋₄ alkyl, C₂₋₄ alkenyl, C₂₋₄ alkynyl or NR₄R₁₄,excluding the moieties SR₅ being SNR₄R₁₄, S(O)₂R₅ being SO₂H and S(O)R₅being SOH;

R₆ is hydrogen, C₁₋₁₀ alkyl, C₃₋₇ cycloalkyl, heterocyclyl, heterocyclylC₁₋₁₀alkyl, aryl, arylC₁₋₁₀ alkyl, heteroaryl or heteroarylC₁₋₁₀ alkyl,wherein these moieties may be optionally substituted;

R₇ is C₁₋₆alkyl, aryl, arylC₁₋₆alkyl, heterocyclic, heterocyclylC₁₋₆alkyl, heteroaryl, or heteroarylC₁₋₆alkyl; and wherein each of thesemoieties may be optionally substituted;

R₈ is hydrogen, C₁₋₄ alkyl, halo-substituted C₁₋₄ alkyl, C₂₋₄ alkenyl,C₂₋₄ alkynyl, C₃₋₇ cycloalkyl, C₅₋₇ cycloalkenyl, aryl, arylC₁₋₄ alkyl,heteroaryl, heteroarylC₁₋₄ alkyl, heterocyclyl, heterocyclylC₁₋₄ alkyl,(CR₁₀R₂₀)_(t)OR₇, (CR₁₀R₂₀)_(t)S(O)_(m)R₇, (CR₁₀R₂₀)_(t)NHS(O)₂R₇, or(CR₁₀R₂₀)_(t)NR₄R₁₄; and wherein the aryl, arylalkyl, heteroaryl,heteroaryl alkyl may be optionally substituted;

R₉ is hydrogen, C(Z)R₆ or optionally substituted C₁₋₁₀ alkyl, optionallysubstituted aryl or optionally substituted aryl-C₁₋₄ alkyl;

R₁₀ and R₂₀ are each independently selected from hydrogen or C₁₋₄ alkyl;

R₁₁ is C₁₋₄ alkyl, halo-substituted C₁₋₄ alkyl, C₂₋₄ alkenyl, C₂₋₄alkynyl, C₃₋₇ cycloalkyl, C₅₋₇ cycloalkenyl, aryl, arylC₁₋₄ alkyl,heteroaryl, heteroarylC₁₋₄ alkyl, heterocyclyl, heterocyclylC₁₋₄ alkyl,(CR₁₀R₂₀)_(t)OR₇, (CR₁₀R₂₀)_(t)S(O)_(m)R₇, (CR₁₀R₂₀)_(t)NHS(O)₂R₇, or(CR₁₀R₂₀)_(v)NR₄R₁₄;

and wherein the aryl, arylalkyl, heteroaryl, heteroaryl alkyl may beoptionally substituted;

Z is oxygen or sulfur;

n is 0, or an integer having a value of 1 to 10;

m is 0, or the integer 1 or 2;

v is 0, or an integer having a value of 1 or 2;

t is an integer having a value of 1 to 3;

or a pharmaceutically acceptable salt thereof.

DETAILED DESCRIPTION OF THE INVENTION

The present invention is directed to novel compounds of Formula (I), orpharmaceutically acceptable salts thereof.

Suitably, for compounds of Formula (I) R₁ is phenyl, naphth-1-yl,naphth-2-yl, heterocyclic or a heteroaryl ring. The R₁ moiety may beoptionally substituted independently by one or more substituents,preferably from one to three substituents each independently selectedfrom halogen, C₁₋₄ alkyl, halo-substituted-C₁₋₄ alkyl, cyano, nitro,(CR₁₀R₂₀)_(v)NR₄R₁₄, (CR₁₀R₂₀)_(v)C(Z)NR₄R₁₄, (CR₁₀R₂₀)_(v)C(Z)OR₈,(CR₁₀R₂₀)_(v)COR₃, (CR₁₀R₂₀)_(v)C(O)H, SR₅, S(O)R₅, S(O)₂R₅,(CR₁₀R₂₀)_(v)OR₈, ZC(Z)R₁₁, NR₁₀C(Z)R₁₁, or NR₁₀S(O)₂R₇.

Preferably, the R₁ moiety is an aryl ring. More preferably, the arylring is a phenyl ring.

Preferably, the R ₁ moiety is substituted by one or more halogens, suchas fluorine or chlorine; an alkyl, hydroxy, alkoxy, amino, orhalosubstituted alkyl, such as CF₃.

Suitably, for compounds of Formula (I), R₂ is an C₁₋₁₀ alkyl, C₃₋₇cycloalkyl, C₃₋₇cycloalkylC₁₋₁₀ alkyl, C₅₋₇ cycloalkenyl, C₅₋₇cycloalkenyl C₁₋₁₀ alkyl, aryl, arylC₁₋₁₀ alkyl, heteroaryl,heteroarylC₁₋₁₀ alkyl, heterocyclic, or heterocyclylC₁₋₁₀ alkyl moiety.These moieties may be optionally substituted one or more times,preferably 1 to 3 times, independently with C₁₋₁₀ alkyl,halo-substituted C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, C₃₋₇cycloalkyl, C₃₋₇cycloalkylC₁₋₁₀ alkyl, C₅₋₇ cycloalkenyl, C₅₋₇cycloalkenyl C₁₋₁₀ alkyl, halogen, (CR₁₀R₂₀)_(n)OR₆, (CR₁₀R₂₀)_(n)SH,(CR₁₀R₂₀)_(n)S(O)_(m)R₇, (CR₁₀R₂₀)_(n)NHS(O)₂R₇, (CR₁₀R₂₀)_(n)NR₄R₁₄,(CR₁₀R₂₀)_(n)CN, (CR₁₀R₂₀)_(n)S(O)₂NR₄R₁₄, (CR₁₀R₂₀)_(n)C(Z)R₆,(CR₁₀R₂₀)_(n)OC(Z)R₆, (CR₁₀R₂₀)_(n)C(Z)OR₆, (CR₁₀R₂₀)_(n)C(Z)NR₄R₁₄,(CR₁₀C₂₀)_(n)NR₁₀C(Z)R₆, (CR₁₀R₂₀)_(n)NR₁₀C(═NR₁₀)NR₄R₁₄,(CR₁₀R₂₀)_(n)OC(Z)NR₄R₁₄, (CR₁₀R₂₀)_(n)NR₁₀C(Z)NR₄R₁₄, or(CR₁₀R₂₀)_(n)NR₁₀C(Z)OR₇.

Preferably, R₂ is an optionally substituted aryl or arylalkyl moiety,such as a unsubstituted or substituted phenyl or benzyl ring; a C₁₋₁₀alkyl, such as isopropyl; C₃₋₇ cycloalkyl, such as cyclohexyl; aheteroarylalkyl, such as an optionally substitued pyrrolidinyl C₁₋₁₀alkyl or pyridyl C₁₋₁₀ alkyl; or an optionally substituted heterocyclicC₁₋₁₀ alkyl, such as a morpholinyl C₁₋₁₀ alkyl. The alkyl portion ofthese chains may as indicated herein be a branced or straight chain.

Preferably the R₂ moiety is substituted by one or more halogens, such asfluorine or chlorine; an alkyl, such as methyl; hydroxy, alkoxy, amino,or a halosubstituted alkyl, such as CF₃.

Preferred R₂ moieties include phenyl, benzyl, phenylethyl, cyclohexyl,isopropyl, optionally substitued pyrrolylethyl, 2,6-difluorphenyl,2-morpholin-4-yl-propyl, 2-morpholin-4-yl-ethyl, or 2-pyridinylethyl.

A preferred substition pattern on the R₂ ring when it is a phenyl orphenylalkyl moiety is in the 2,6-position.

Suitably, n is 0, or an integer having a value of 1 to 10.

Suitably, m is 0, or the integer 1 or 2.

Suitably, v is 0, or an integer having a value of 1 or 2.

Suitably, t is an integer having a value of 1 to 3.

Suitably, Z is oxygen or sulfur, preferably oxygen.

Suitably, R₃ is C₁₋₄ alkyl, halo-substituted C₁₋₄ alkyl, C₂₋₄ alkenyl,C₂₋₄ alkynyl, C₃₋₇ cycloalkyl, C₅₋₇ cycloalkenyl, aryl, arylC₁₋₄ alkyl,heteroaryl, heteroarylC₁₋₄ alkyl, heterocyclyl, heterocyclylC₁₋₄ alkyl,(CR₁₀R₂₀)_(v)OR₇, (CR₁₀R₂₀)_(v)S(O)_(m)R₇, (CR₁₀R₂₀)_(v)NHS(O)₂R₇, or(CR₁₀R₂₀)_(v)NR₄R₁₄; and wherein the aryl, arylalkyl, heteroaryl,heteroaryl alkyl may be optionally substituted.

Suitably, R₄ and R₁₄ is each independently selected from hydrogen oroptionally substituted C₁₋₄ alkyl, optionally substituted aryl oroptionally substituted aryl-C₁₋₄ alkyl, or together with the nitrogenwhich they are attached form a heterocyclic ring of 5 to 7 members whichring optionally contains an additional heteroatom selected from oxygen,sulfur or NR₉.

Suitably, R₅ is hydrogen, C₁₋₄ alkyl, C₂₋₄ alkenyl, C₂₋₄ alkynyl orNR₄R₁₄, excluding the moieties SR₅ being SNR₄R₁₄, S(O)₂R₅ being SO₂H andS(O)R₅ being SOH.

Suitably, R₆ is hydrogen, C₁₋₁₀ alkyl, C₃₋₇ cycloalkyl, heterocyclyl,heterocyclyl C₁₋₁₀ alkyl, aryl, arylC₁₋₁₀ alkyl, heteroaryl orheteroarylC₁₋₁₀ alkyl, wherein these moieties may be optionallysubstituted.

Suitably, R₇ is C₁₋₆alkyl, aryl, arylC₁₋₆alkyl, heterocyclic,heterocyclylC₁₋₆ alkyl, heteroaryl, or heteroarylC₁₋₆alkyl; and whereineach of these moieties may be optionally substituted.

Suitably, R₈ is hydrogen, C₁₋₄ alkyl, halo-substituted C₁₋₄ alkyl, C₂₋₄alkenyl, C₂₋₄ alkynyl, C₃₋₇ cycloalkyl, C₅₋₇ cycloalkenyl, aryl,arylC₁₋₄ alkyl, heteroaryl, heteroarylC₁₋₄ alkyl, heterocyclyl,heterocyclylC₁₋₄ alkyl, (CR₁₀R₂₀)_(t)OR₇, (CR₁₀R₂₀)_(t)S(O)_(m)R₇,(CR₁₀R₂₀)_(t)NHS(O)₂R₇, or (CR₁₀R₂₀)_(t)NR₄R₁₄; and wherein the aryl,arylalkyl, heteroaryl, heteroaryl alkyl may be optionally substituted.

Suitably, R₉ is hydrogen, C(Z)R₆ or optionally substituted C₁₋₁₀ alkyl,optionally substituted aryl or optionally substituted aryl-C₁₋₄ alkyl.

Suitably, R₁₀ and R₂₀ are each independently selected from hydrogen orC₁₋₄ alkyl.

Suitably, R₁₁ is C₁₋₄ alkyl, halo-substituted C₁ ₄ alkyl, C₂₋₄ alkenyl,C₂₋₄ alkynyl, C₃₋₇ cycloalkyl, C₅₋₇ cycloalkenyl, aryl, arylC₁₋₄ alkyl,heteroaryl, heteroarylC₁₋₄ alkyl, heterocyclyl, heterocyclylC₁₋₄ alkyl,(CR₁₀R₂₀)_(t)OR₇, (CR₁₀R₂₀)_(t)S(O)_(m)R₇, (CR₁₀R₂₀)_(t)NHS(O)₂R₇, or(CR₁₀R₂₀)_(v)NR₄R₁₄; and wherein the aryl, arylalkyl, heteroaryl,heteroaryl alkyl may be optionally substituted.

As used herein, “optionally substituted” unless specifically definedshall mean such groups as halogen, such as fluorine, chlorine, bromineor iodine; hydroxy; hydroxy substituted C₁₋₁₀alkyl; C₁₋₁₀ alkoxy, suchas methoxy or ethoxy; halosubstituted C₁₋₁₀ alkoxy; S(O)m alkyl, such asmethyl thio, methylsulfinyl or methyl sulfonyl; NR₇R₁₇, such as amino ormono or -disubstituted C₁₋₄ alkyl or wherein the R₇R₁₇ can cyclizetogether with the nitrogen to which they are attached to form a 5 to 7membered ring which optionally contains an additional heteroatomselected from O/N/S; C₁₋₁₀ alkyl, C₃₋₇cycloalkyl, or C₃₋₇cycloalkylC₁₋₁₀ alkyl group, such as methyl, ethyl, propyl, isopropyl, t-butyl,etc. or cyclopropyl methyl; halosubstituted C₁₋₁₀ alkyl, such CF₂CF₂H,or CF₃; an optionally substituted aryl, such as phenyl, or an optionallysubstituted arylalkyl, such as benzyl or phenethyl, wherein these arylcontaining moieties may also be substituted one to two times by halogen;hydroxy; hydroxy substituted alkyl; C₁₋₁₀ alkoxy; S(O)_(m)alkyl; amino,mono & di-substituted C₁₋₄ alkyl amino, such as in the NR₇R₁₇ group;C₁₋₄ alkyl, or CF₃.

Suitable pharmaceutically acceptable salts are well known to thoseskilled in the art and include basic salts of inorganic and organicacids, such as hydrochloric acid, hydrobromic acid, sulphuric acid,phosphoric acid, methane sulphonic acid, ethane sulphonic acid, aceticacid, malic acid, tartaric acid, citric acid, lactic acid, oxalic acid,succinic acid, fumaric acid, maleic acid, benzoic acid, salicylic acid,phenylacetic acid and mandelic acid.

In addition, pharmaceutically acceptable salts of compounds of Formula(I) may also be formed with a pharmaceutically acceptable cation, forinstance, if a substituent group comprises a carboxy moiety. Suitablepharmaceutically acceptable cations are well known to those skilled inthe art and include alkaline, alkaline earth, ammonium and quaternaryammonium cations.

The term “halo” or “halogens” is used herein to mean the halogens,chloro, fluoro, bromo and iodo.

The term “C₁₋₁₀alkyl” or “alkyl” or “alkyl₁₋₁₀” is used herein to meanboth straight and branched chain radicals of 1 to 10 carbon atoms,unless the chain length is otherwise limited, including, but not limitedto, methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, iso-butyl,tert-butyl, n-pentyl and the like.

The term “cycloalkyl” is used herein to mean cyclic radicals, preferablyof 3 to 8 carbons, including but not limited to cyclopropyl,cyclopentyl, cyclohexyl, and the like.

The term “cycloalkenyl” is used herein to mean cyclic radicals,preferably of 5 to 8 carbons, which have at least one bond including butnot limited to cyclopentenyl, cyclohexenyl, and the like.

The term “alkenyl” is used herein at all occurrences to mean straight orbranched chain radical of 2-10 carbon atoms, unless the chain length islimited thereto, including, but not limited to ethenyl, 1-propenyl,2-propenyl, 2-methyl-1-propenyl, 1-butenyl, 2-butenyl and the like.

The term “aryl” is used herein to mean phenyl and naphthyl.

The term “heteroaryl” (on its own or in any combination, such as“heteroaryloxy”, or “heteroaryl alkyl”) is used herein to mean a 5-10membered aromatic ring system in which one or more rings contain one ormore heteroatoms selected from the group consisting of N, O or S, suchas, but not limited, to

The term “heterocyclic” (on its own or in any combination, such as“heterocyclylalkyl”) is used herein to mean a saturated or partiallyunsaturated 4-10 membered ring system in which one or more rings containone or more heteroatoms selected from the group consisting of N, O, orS, such as, but not limited to, pyrrolidine, piperidine, piperazine,morpholine, tetrahydropyran, imidazolidine, or pyrazolidine.

The term “aralkyl” or “heteroarylalkyl” or “heterocyclicalkyl” is usedherein to mean C₁₋₄ alkyl as defined above attached to an aryl,heteroaryl or heterocyclic moiety as also defined herein unlessotherwise indicate.

The term “sulfinyl” is used herein to mean the oxide S (O) of thecorresponding sulfide, the term “thio” refers to the sulfide, and theterm “sulfonyl” refers to the fully oxidized S (O)₂ moiety.

The term “aroyl” is used herein to mean C(O)Ar, wherein Ar is as phenyl,naphthyl, or aryl alkyl derivative such as defined above, such groupinclude but are not limited to benzyl and phenethyl.

The term “alkanoyl” is used herein to mean C(O)C₁₋₁₀ alkyl wherein thealkyl is as defined above.

It is recognized that the compounds of the present invention may existas stereoisomers, regioisomers, or diastereiomers. These compounds maycontain one or more asymmetric carbon atoms and may exist in racemic andoptically active forms. All of these compounds are included within thescope of the present invention.

Exemplified compounds of Formula (I), according to the invention includethose mentioned in the Examples and their pharmaceutically acceptablesalts.

The compounds of Formula (I) may be obtained by applying syntheticprocedures, described herein. The synthesis provided for is applicableto producing compounds of Formula (I) having a variety of different R₁,and R₂, groups which are reacted, employing optional substituents whichare suitably protected, to achieve compatibility with the reactionsoutlined herein. Subsequent deprotection, in those cases, then affordscompounds of the nature generally disclosed.

Once the nucleus has been established, further compounds of Formula (I)may be prepared by applying standard techniques for functional groupinterconversion, well known in the art. For instance: C(O)NR₄R₁₄ fromCO₂CH₃ by heating with or without catalytic metal cyanide, e.g. NaCN,and HNR₄R₁₄ in CH₃OH; OC(O)R₃ from OH with e.g., ClC(O)R₃ in pyridine;NR₁₀—C(S)NR₄R₁₄ from NHR₁₀ with an alkylisothiocyante or thiocyanicacid; NR₁₀C(O)OR₇ from NHR₁₀ with the alkyl chloroformate;NR₁₀C(O)NR₄R₁₄ from NHR₁₀ by treatment with an isocyanate, e.g. HN═C═Oor R₁₀N═C═O; NR₁₀—C(O)R₇ from NHR₁₀ by treatment with Cl—C(O)R₇ inpyridine; C(═NR₁₀)NR₄R₁₄ from C(NR₄R₁₄)SR₃ with H₃NR₃ ⁺OAc³¹ by heatingin alcohol; C(NR₄R₁₄)SR₃ from C(S)NR₄R₁₄ with R₆-I in an inert solvent,e.g. acetone; C(S)NR₄R₁₄ (where R₄ or R₁₄ is not hydrogen) from C(S)NH₂with HNR₄R₁₄—C(═NCN)—NR₄R₁₄ from C(═NR₄R₁₄)—SR₃ with NH₂CN by heating inanhydrous alcohol, alternatively from C(═NH)—NR₄R₁₄ by treatment withBrCN and NaOEt in EtOH; NR₁₀—C(═NCN)SR₈ from NHR₁₀ by treatment with(R₈S)2C═NCN; NR₁₀SO₂R₃ from NHR₁₀ by treatment with ClSO₂R₃ by heatingin pyridine; NR₁₀C(S)R₆ from NR₁₀C(O)R₆ by treatment with Lawesson'sreagent[2,4-bis(4-methoxyphenyl)-1,3,2,4-dithiadiphosphetane-2,4-disulfide];NR₁₀SO₂CF₃ from NHR₆ with triflic anhydride and base wherein R₃, R₆, R₇,R₁₀, R₄ and R₁₄ are as defined in Formula (I) herein.

Precursors of the groups R₁, and R₂ can be other R₁, and R₂ groups whichcan be interconverted by applying standard techniques for functionalgroup interconversion. For example wherein a moiety is a halosubstituted C₁₋₁₀ alkyl can be converted to the corresponding C₁₋₁₀alkylN₃ derivative by reacting with a suitable azide salt, andthereafter if desired can be reduced to the corresponding C₁₋₁₀alkylNH₂compound, which in turn can be reacted with R₇S(0)₂X wherein X is halo(e.g., chloro) to yield the corresponding C₁₋₁₀alkylNHS(0)₂R₇ compound.

Alternatively wherein the moiety is a halo-substituted C₁₋₁₀-alkyl itcan be reacted with an amine R₄R₁₄NH to yield the correspondingC₁₋₁₀-alkylNR₄R₁₄ compound, or can be reacted with an alkali metal saltof R₇SH to yield the corresponding C₁₋₁₀alkylSR₇ compound.

Suitable protecting groups for use with hydroxyl groups and nitrogengroups are well known in the art and described in many references, forinstance, Protecting Groups in Organic Synthesis, Greene T W,Wiley-Interscience, New York, 1981. Suitable examples of hydroxylprotecting groups include silyl ethers, such as t-butyldimethyl ort-butyldiphenyl, and alkyl ethers, such as methyl connected by an alkylchain of variable link, (CR₁₀R₂₀)_(n).

Pharmaceutically acid addition salts of compounds of Formula (I) may beobtained in known manner, for example by treatment thereof with anappropriate amount of acid in the presence of a suitable solvent.

For purposes herein in Scheme 1, Ra and Rb are as defined in Formula (I)as R₂ and R₁ respectively.

Synthesis of 3,4-Dihydro-(1H)-quinazolin-2-ones substituted with an arylor heteroaryl substituent in the 5 position (Rb) can be prepared bystarting from a suitable 2,3 disubstituted benzonitrile (1). Some ofthese benzonitriles are commercially available while others can beprepared from commercially available 2-amino-6-halobenzonitrilesfollowing literature procedures such as that exemplified by Hynes, JohnB. et. al., J. Hetercycl. Chem. 1988, 25, 1173-1177. Reaction of (1)with a boronic acid or derivative under typical aryl coupling conditionssuch as the Suzuki reaction described in Miyaura and Suzuki, Chem. Rev.1995, 95, 2547-2483 gives the cross-coupling product (2). Many of therequired boronic acids or their derivatives are commercially availableand others can be prepared by methods outlined in Miyaura and Suzuki,Chem. Rev. 1995, 95, 2547-2483. The cross coupled products (2) can alsobe prepared by a Stille cross-coupling reaction as outlined in Mitchell,T. N., Synthesis 1992, 803-815. Reaction of coupled product (2) with anappropriate primary amine, aniline, or heteroaryl amine, many of whichare commercially available, in a suitable solvent such as dimethylsulfoxide or dimethylformamide will give the secondary amine (3). Incases of aliphatic amines, the amine should generally have sufficientnucleophilicity to displace the halide without addition of a strongerbase to form the amine anion as demonstrated in Hynes, John B. et. al.,J. Hetercycl. Chem. 1988, 25, 1173-1177. In the case of the aromaticamines such as aniline, preforming the anion by the use of a strong basesuch as sodium hydride or potassium t-butoxide should facilitate thereaction as to exemplified in Mettey, Yvette et. al., Heterocycles,1993, 36, 987-993, and Govin, John H., J. Chem. Soc., Perkin Trans. 1,1988, 6, 1331-1335. Reduction of the nitrile (3) to give the benzylamine (4) can be accomplished with a variety of reducing agents such aslithium aluminum hydride in a suitable solvent such as diethyl ether ortetrahydrofuran at temperatures ranging from 0° C. to reflux. Reductionof the nitrile to the amine may also be carried out by hydrogenationover a variety of catalysts such as Raney nickel, platinum or palladiumin a suitable solvent such as ethanol, methanol, ethyl acetate, oracetic acid. Alternatively the reduction could be carried out using anyof the appropriate boranes or borohydride reducing agents in a suitablesolvent such as tetrahydrofuran or diethyl ether. Cyclization of thediamino compound (4) to the desired 3,4-dihydro-(1H)-quinazolin-2-ones(5), can be accomplished by a variety of synthetic procedures only a fewof which are exemplified here. Cyclization can be accomplished using1,1′-carbonyldiimidazole as demonstrated in Takai, Haruki et. al., Chem.Pharm. Bull., 1985, 33, 1116-1128. Cyclization can also be carried outwith phosgene as demonstrated by Szabo, Janos, et. al., J. Heterocycl.Chem. 1992, 29, 1513-1517. Alternatively, cyclization can be carried outusing chloroformates as exemplified in Kornet, Milton, J. Heterocycl.Chem. 1992,29, 103-105.

Methods of Treatment

The compounds of Formula (I) or a pharmaceutically acceptable saltthereof can be used in the manufacture of a medicament for theprophylactic or therapeutic treatment of any disease state in a human,or other mammal, which is exacerbated or caused by excessive orunregulated cytokine production by such mammal's cell, such as but notlimited to monocytes and/or macrophages.

Compounds of Formula (I) are capable of inhibiting proinflammatorycytokines, such as IL-1, IL-6, IL-8, and TNF and are therefore of use intherapy. IL-1, IL-6, IL-8 and TNF affect a wide variety of cells andtissues and these cytokines, as well as other leukocyte-derivedcytokines, are important and critical inflammatory mediators of a widevariety of disease states and conditions. The inhibition of thesepro-inflammatory cytokines is of benefit in controlling, reducing andalleviating many of these disease states.

Accordingly, the present invention provides a method of treating acytokine-mediated disease which comprises administering an effectivecytokine-interfering amount of a compound of Formula (I) or apharmaceutically acceptable salt thereof.

Compounds of Formula (I) are capable of inhibiting inducibleproinflammatory proteins, such as COX-2, also referred to by many othernames such as prostaglandin endoperoxide synthase-2 (PGHS-2) and aretherefore of use in therapy. These proinflammatory lipid mediators ofthe cyclooxygenase (CO) pathway are produced by the inducible COX-2enzyme. Regulation, therefore of COX-2 which is responsible for thethese products derived from arachidonic acid, such as prostaglandinsaffect a wide variety of cells and tissues are important and criticalinflammatory mediators of a wide variety of disease states andconditions. Expression of COX-1 is not effected by compounds of Formula(I). This selective inhibition of COX-2 may alleviate or spareulcerogenic liability associated with inhibition of COX-1 therebyinhibiting prostoglandins essential for cytoprotective effects. Thusinhibition of these pro-inflammatory mediators is of benefit incontrolling, reducing and alleviating many of these disease states. Mostnotably these inflammatory mediators, in particular prostaglandins, havebeen implicated in pain, such as in the sensitization of pain receptors,or edema. This aspect of pain management therefore includes treatment ofneuromuscular pain, headache, cancer pain, and arthritis pain. Compoundsof Formula (I) or a pharmaceutically acceptable salt thereof, are of usein the prophylaxis or therapy in a human, or other mammal, by inhibitionof the synthesis of the COX-2 enzyme.

Accordingly, the present invention provides a method of inhibiting thesynthesis of COX-2 which comprises administering an effective amount ofa compound of Formula (I) or a pharmaceutically acceptable salt thereof.The present invention also provides for a method of prophylaxistreatment in a human, or other mammal, by inhibition of the synthesis ofthe COX-2 enzyme.

In particular, compounds of Formula (I) or a pharmaceutically acceptablesalt thereof are of use in the prophylaxis or therapy of any diseasestate in a human, or other mammal, which is exacerbated by or caused byexcessive or unregulated IL-1, IL-6, IL-8 or TNF production by suchmammal's cell, such as, but not limited to, monocytes and/ormacrophages.

Accordingly, in another aspect, this invention relates to a method ofinhibiting the production of IL-1 in a mammal in need thereof whichcomprises administering to said mammal an effective amount of a compoundof Formula (I) or a pharmaceutically acceptable salt thereof.

There are many disease states in which excessive or unregulated IL-1production is implicated in exacerbating and/or causing the disease.These include rheumatoid arthritis, osteoarthritis, meningitis, ischemicand hemorrhagic stroke, neurotrauma/closed head injury, stroke,endotoxemia and/or toxic shock syndrome, other acute or chronicinflammatory disease states such as the inflammatory reaction induced byendotoxin or inflammatory bowel disease, tuberculosis, atherosclerosis,muscle degeneration, multiple sclerosis, cachexia, bone resorption,psoriatic. arthritis, Reiter's syndrome, rheumatoid arthritis, gout,traumatic arthritis, rubella arthritis and acute synovitis. Recentevidence also links IL-1 activity to diabetes, pancreatic β celldiseases and Alzheimer's disease.

Use of a CSAID for the treatment of CSBP mediated disease states, caninclude, but not be limited to neurodegenerative diseases, such asAlzheimer's disease (as noted above), Parkinson's disease and multiplesclerosis, etc.

In a further aspect, this invention relates to a method of inhibitingthe production of TNF in a mammal in need thereof which comprisesadministering to said mammal an effective amount of a compound ofFormula (I) or a pharmaceutically acceptable salt thereof.

Excessive or unregulated TNF production has been implicated in mediatingor exacerbating a number of diseases including rheumatoid arthritis,rheumatoid spondylitis, osteoarthritis, gouty arthritis and otherarthritic conditions, sepsis, septic shock, endotoxic shock, gramnegative sepsis, toxic shock syndrome, adult respiratory distresssyndrome, chronic pulmonary inflammatory disease and chronic obstructivepulmonary disease, silicosis, pulmonary sarcoisosis, bone resorptiondiseases, such as osteoporosis, cardiac, brain and renal reperfusioninjury, graft vs. host reaction, allograft rejections, fever andmyalgias due to infection, such as influenza, brain infections includingencephalitis (including HIV-induced forms), cerebral malaria,meningitis, ischemic and hemorrhagic stroke, cachexia secondary toinfection or malignancy, cachexia secondary to acquired immunedeficiency syndrome (AIDS), AIDS, ARC (AIDS related complex), keloidformation, scar tissue formation, inflammatory bowel disease, Crohn'sdisease, ulcerative colitis and pyresis.

Compounds of Formula (I) are also useful in the treatment of viralinfections, where such viruses are sensitive to upregulation by TNF orwill elicit TNF production in vivo. The viruses contemplated fortreatment herein are those that produce TNF as a result of infection, orthose which are sensitive to inhibition, such as by decreasedreplication, directly or indirectly, by the TNF inhibiting-compounds ofFormula (I). Such viruses include, but are not limited to HIV-1, HIV-2and HIV-3, Cytomegalovirus (CMV), Influenza, adenovirus and the Herpesgroup of viruses, such as but not limited to, Herpes Zoster and HerpesSimplex. Accordingly, in a further aspect, this invention relates to amethod of treating a mammal afflicted with a human immunodeficiencyvirus (HIV) which comprises administering to such mammal an effectiveTNF inhibiting amount of a compound of Formula (I) or a pharmaceuticallyacceptable salt thereof.

It is also recognized that both IL-6 and IL-8 are produced duringrhinovirus (HRV) infections and contribute to the pathogenesis of commoncold and exacerbation of asthma associated with HRV infection (Turner etal. (1998), Clin. Infec. Dis., Vol 26, p 840; Teren et al. (1997), Am JRespir Crit Care Med vol 155, p1362; Grunberg et al. (1997), Am J RespirCrit Care Med 156:609 and Zhu et al, J Clin Invest (1996), 97:421). Ithas also been demonstrated in vitro that infection of pulmonaryepithelial cells with HRV results in production of IL-6 and IL-8(Subauste et al., J. Clin. Invest. 1995, 96:549.) Epithelial cellsrepresent the primary site of infection of HRV. Therefore another aspectof the present invention is a method of treatment to reduce inflammationassociated with a rhinovirus infection, not necessarily a direct effecton virus itself.

Compounds of Formula (I) may also be used in association with the,veterinary treatment of mammals, other than in humans, in need ofinhibition of TNF production. TNF mediated diseases for treatment,therapeutically or prophylactically, in animals include disease statessuch as those noted above, but in particular viral infections. Examplesof such viruses include, but are not limited to, lentivirus infectionssuch as, equine infectious anaemia virus, caprine arthritis virus, visnavirus, or maedi virus or retrovirus infections, such as but not limitedto feline immunodeficiency virus (FIV), bovine immunodeficiency virus,or canine immunodeficiency virus or other retroviral infections.

The compounds of Formula (I) may also be used topically in the treatmentor prophylaxis of topical disease states mediated by or exacerbated byexcessive cytokine production, such as by IL-1 or TNF respectively, suchas inflamed joints, eczema, psoriasis and other inflammatory skinconditions such as sunburn; inflammatory eye conditions includingconjunctivitis; pyresis, pain and other conditions associated withinflammation. Periodontal disease has also been implemented in cytokineproduction, both topically and systemically. Hence use of compounds ofFormula (I) to control the inflammation associated with cytokineproduction in such peroral diseases such as gingivitis and periodontitisis another aspect of the present invention.

Compounds of Formula (I) have also been shown to inhibit the productionof IL-8 (Interleukin-8, NAP). Accordingly, in a further aspect, thisinvention relates to a method of inhibiting the production of IL-8 in amammal in need thereof which comprises administering to said mammal aneffective amount of a compound of Formula (I) or a pharmaceuticallyacceptable salt thereof.

There are many disease states in which excessive or unregulated IL-8production is implicated in exacerbating and/or causing the disease.These diseases are characterized by massive neutrophil infiltration suchas, psoriasis, inflammatory bowel disease, asthma, cardiac, brain andrenal reperfusion injury, adult respiratory distress syndrome,thrombosis and glomerulonephritis. All of these diseases are associatedwith increased IL-8 production which is responsible for the chemotaxisof neutrophils into the inflammatory site. In contrast to otherinflammatory cytokines (IL-1, TNF, and IL-6), IL-8 has the uniqueproperty of promoting neutrophil chemotaxis and activation. Therefore,the inhibition of IL-8 production would lead to a direct reduction inthe neutrophil infiltration.

The compounds of Formula (I) are administered in an amount sufficient toinhibit cytokine, in particular IL-1, IL-6, IL-8 or TNF, production suchthat it is regulated down to normal levels, or in some case to subnormallevels, so as to ameliorate or prevent the disease state. Abnormallevels of IL-1, IL-6, IL-8; or TNF, for instance in the context of thepresent invention, constitute: (i) levels of free (not cell bound) IL-1,IL-6, IL-8 or TNF greater than or equal to 1 picogram per ml; (ii) anycell associated IL-1, IL-6, IL-8 or TNF; or (iii) the presence of IL-1,IL-6, IL-8 or TNF mRNA above basal levels in cells or tissues in whichIL-1, IL-6, IL-8 or TNF, respectively, is produced.

The discovery that the compounds of Formula (I) are inhibitors ofcytokines, specifically IL-1, IL-6, IL-8 and TNF is based upon theeffects of the compounds of Formulas (I) on the production of the IL-1,IL-8 and TNF in in vitro assays which are described herein.

As used herein, the term “inhibiting the production of IL-1 (IL-6, IL-8or TNF)” refers to:

a) a decrease of excessive in vivo levels of the cytokine (IL-1, IL-6,IL-8 or TNF) in a human to normal or sub-normal levels by inhibition ofthe in release of the cytokine by all cells, including but not limitedto monocytes or macrophages;

b) a down regulation, at the genomic level, of excessive in vivo levelsof the cytokine (IL-1, IL-6, IL-8 or TNF) in a human to normal orsub-normal levels;

c) a down regulation, by inhibition of the direct synthesis of thecytokine (IL-1, IL-6, IL-8 or TNF) as a postranslational event; or

d) a down regulation, at the translational level, of excessive in vivolevels of the cytokine (IL-1, IL-6, IL-8 or TNF) in a human to normal orsub-normal levels.

As used herein, the term “TNF mediated disease or disease state” refersto any and all disease states in which TNF plays a role, either byproduction of TNF itself, or by TNF causing another monokine to bereleased, such as but not limited to IL-1, IL-6 or IL-8. A disease statein which, for instance, IL-1 is a major component, and whose productionor action, is exacerbated or secreted in response to TNF, wouldtherefore be considered a disease stated mediated by TNF.

As used herein, the term “cytokine” refers to any secreted polypeptidethat affects the functions of cells and is a molecule which modulatesinteractions between cells in the immune, inflammatory or hematopoieticresponse. A cytokine includes, but is not limited to, monokines andlymphokines, regardless of which cells produce them. For instance, amonokine is generally referred to as being produced and secreted by amononuclear cell, such as a macrophage and/or monocyte. Many other cellshowever also produce monokines, such as natural killer cells,fibroblasts, basophils, neutrophils, endothelial cells, brainastrocytes, bone marrow stromal cells, epideral keratinocytes andB-lymphocytes. Lymphokines are generally referred to as being producedby lymphocyte cells. Examples of cytokines include, but are not limitedto, Interleukin-1 (IL-1), Interleukin-6 (IL-6), Interleukin-8 (IL-8),Tumor Necrosis Factor-alpha (TNF-α) and Tumor Necrosis Factor beta(TNF-β).

As used herein, the term “cytokine interfering” or “cytokine suppressiveamount” refers to an effective amount of a compound of Formula (I) whichwill cause a decrease in the in vivo levels of the cytokine to normal orsub-normal levels, when given to a patient for the prophylaxis ortreatment of a disease state which is exacerbated by, or caused by,excessive or unregulated cytokine production.

As used herein, the cytokine referred to in the phrase “inhibition of acytokine, for use in the treatment of a HIV-infected human” is acytokine which is implicated in

(a) the initiation and/or maintenance of T cell activation and/oractivated T cell-mediated HIV gene expression and/or replication and/or(b) any cytokine-mediated disease associated problem such as cachexia ormuscle degeneration.

As TNF-β (also known as lymphotoxin) has close structural homology withTNF-α (also known as cachectin) and since each induces similar biologicresponses and binds to the same cellular receptor, both TNF-α and TNF-βare inhibited by the compounds of the present invention and thus areherein referred to collectively as “TNF” unless specifically delineatedotherwise.

A new member of the MAP kinase family, alternatively termed CSBP, p38,or RK, has been identified independently by several laboratories.Activation of this novel protein kinase via dual phosphorylation hasbeen observed in different cell systems upon stimulation by a widespectrum of stimuli, such as physicochemical stress and treatment withlipopolysaccharide or proinflammatory cytokines such as interleukin-1and tumor necrosis factor. The cytokine biosynthesis inhibitors, of thepresent invention, compounds of Formula (I) have been determined to bepotent and selective inhibitors of CSBP/p38/RK kinase activity. Theseinhibitors are of aid in determining the signaling pathway involvementin inflammatory responses. In particular, for the first time adefinitive signal transduction pathway can be prescribed to the actionof lipopolysaccharide in cytokine production in macrophages. In additionto those diseases already noted, treatment of stroke, neurotrauma,cardiac and renal reperfusion injury, congestive heart failure, chronicrenal failure, angiogenesis & related processes, such as cancer,thrombosis, glomerulonephritis, diabetes and pancreatic b cells,multiple sclerosis, muscle degeneration , eczema, psoriasis, sunburn,and conjunctivitis are also included.

The CSBP inhibitors were subsequently tested in a number of animalmodels for anti-inflammatory activity. Model systems were chosen thatwere relatively insensitive to cyclooxygenase inhibitors in order toreveal the unique activities of cytokine suppressive agents. Theinhibitors exhibited significant activity in many such in vivo studies.Most notable are its effectiveness in the collagen-induced arthritismodel and inhibition of TNF production in the endotoxic shock model. Inthe latter study, the reduction in plasma level of TNF correlated withsurvival and protection from endotoxic shock related mortality. Also ofgreat importance are the compounds effectiveness in inhibiting boneresorption in a rat fetal long bone organ culture system. Griswold etal., (1988) Arthritis Rheum. 31:1406-1412; Badger, et al., (1989) Circ.Shock 27, 51-61; Votta et al., (1994)in vitro. Bone 15, 533-538; Lee etal., (1993). B Ann. N. Y. Acad. Sci. 696, 149-170.

Chronic diseases which have an inappropriate angiogenic component arevarious ocular neovasularizations, such as diabetic retinopathy andmacular degeneration. Other chronic diseases which have an excessive orincreased proliferation of vasculature are tumor growth and metastasis,atherosclerosis, and certain arthritic conditions. Therefore CSBP kinaseinhibitors will be of utility in the blocking of the angiogeniccomponent of these disease states.

The term “excessive or increased proliferation of vasculatureinappropriate angiogenesis” as used herein includes, but is not limitedto, diseases which are characterized by hemangiomas and ocular diseases.

The term “inappropriate angiogenesis” as used herein includes, but isnot limited to, diseases which are characterized by vesicleproliferation with accompanying tissue proliferation, such as occurs incancer, metastasis, arthritis and atherosclerosis.

Another aspect of the present invention is directed to the treatment ofinhaled smoke induced airway inflammation, lung chemokine production andcytokine production. The invention may be directed to treatment of theairway induced inflammation which is secondary to other respiratorydisorders such as viral infections that exacerbate asthma (induced bysuch infections), chronic bronchitis, chronic obstructive pulmonarydisease, otitis media, and sinusitis. A respiratory viral infectiontreated in conjunction with the smoke related airway inflammation mayalso be associated with a secondary bacterial infection, such as otitismedia, sinusitis, or pneumonia.

It is noted that the inflammation may be due to cytokines and chemokinerelease from neutrophile activation and other leukocytes, as well asvascular and airway endothelial cell activiation.

For use herein treatment may include prophylaxis for use in a treatmentgroup who may be susceptible to such airway inflammation. It may alsoinclude reducing the symptoms of, ameliorating the symptoms of, reducingthe severity of, reducing the incidence of, or any other change in thecondition of the patient, which improves the therapeutic outcome.

Suitable patient populations for whom this may be prophylaticallybeneficial could be firemen who routinely inhale smoke in the course oftheir duties; use in the military, and by civilians in wartime exposure.

As noted, smoke of natural causes, such as plant extracts, naturalplants products, synthetic material, chemically treated naturalmaterials, or natural products such as oil and gas or other fossilfuels, may be treated within the scope of this invention. Suitably, thetreatment including prophylaxis is related to cigarette smoke orsynthetic/composites, such as occur in fires associated with buringbuildings or homes.

Another aspect of the present invention relates to the use of a CSBP/p38kinase inhibitor for the treatment, including prophylaxis, of thehypertussive activity associated withwith resulting airway inflammationand/or cough in a mammal in need thereof.

The present invention also relates to use of a CSBP/p38 kinase inhibitorfor the treatment, including prophylaxis, of the inflammation enhancedcought related disorders in a mammal in need thereof.

The present invention is also directed to the use of a compound ofFormula (I) in eosinophilic bronchitis, and in cough variant asthma.

The compounds of Formula (I) may also be used in the treatment,including prophylaxis, of eosinophilic inflammation in the airways andcough. Treatment, including prophylaxis is appropriate for eosinophilicbronchitis (as this differs from asthma) and for the treatment,including prophylaxis of cough variant asthma. These disorders may bedirected to treatment of the airway induced inflammation which issecondary to other respiratory disorders such as viral infections thatexacerbate asthma (induced by such infections), chronic bronchitis,chronic obstructive: pulmonary disease, otitis media, and sinusitis. Arespiratory viral infection treated in conjunction with the smokerelated airway inflammation may also be associated with a secondarybacterial infection, such as otitis media, sinusitis, or pneumonia.

The hypertussive or inflammation enhanced cough related disorders mayeither be a direct result of or an association with eosinophiliaactivity. It may also be a result of, or associated with the blockingproduction of certain cytokines which may mediate these phenomena.

For use herein treatment may include prophylaxis for use in a treatmentgroup who may be susceptible to such airway inflammation, and/or cough.It may also include reducing the symptoms of, ameliorating the symptomsof, reducing the severity of, reducing the incidence of, or any otherchange in the condition of the patient, which improves the therapeuticoutcome.

Clinically, eosinophilic bronchitis presents as chronic cough and sputumeosinophilia, but without the abnormalities of airway function seen inasthma. In contrast to cough in patients without sputum eosinophilia,the cough responds to anti-inflammatory therapy, such as inhaledcorticosteroids (Niimi et al., Eosinophilic inflammation in coughvariant asthma, European Respiratory Journal. 11 (5): 1064-9, (1998)).

Patients with cough-variant asthma may also have the following criteria:(1) have not been previously diagnosed as having asthma; (2) complain ofa cough of at least a 3-week duration; (3) do not complain of wheezing,shortness of breath, or chest tightness; (4) have normal results ofphysical examinations; (5) have normal or nearly normal results ofspirometry; (6) have evidence of bronchial hyperresponsiveness duringbronchoprovocation challenge testing; and (7) have a favorable responseto asthma medications (Irwin et al., Interpretation of positive resultsof a methacholine inhalation challenge and 1 week of inhaledbronchodilator use in diagnosing and treating cough-variant asthma(Archives of Internal Medicine. 157(17):1981-1987, (1997)).

Unlike conventional anti-tussive agents, such as codeine ordextromethorphan, a p38 kinase inhibitor appears to have no directantitussive activity, but reduces the airway eosinophilia and normalizesthe hypertussive state. Therefore, use of a p38 inhibitor will reducethe added coughs, or hypertussive state, back to a normal level whichcan be suitably treated with conventional agents and/or therapies asappropriate. Use of the p38 inhibitors will allow for the maintenance ofpatients who are subject to increased cough responsiveness, especiallyunproductive cough, due to other underlying disorders or treatments.This increased cough responsiveness may be modulated, or decreased byuse of this innovative anti-inflammatory therapy.

Accordingly, the present invention provides a method of treating a CSBPkinase mediated disease in a mammal in need thereof, preferably a human,which comprises administering to said mammal, an effective amount of acompound of Formula (I) or a pharmaceutically acceptable salt thereof.

In order to use a compound of Formula (I) or a pharmaceuticallyacceptable salt thereof in therapy, it will normally be formulated intoa pharmaceutical composition in accordance with standard pharmaceuticalpractice. This invention, therefore, also relates to a pharmaceuticalcomposition comprising an effective, non-toxic amount of a compound ofFormula (I) and a pharmaceutically acceptable carrier or diluent.

Compounds of Formula (I), pharmaceutically acceptable salts thereof andpharmaceutical compositions incorporating such may conveniently beadministered by any of the routes conventionally used for drugadministration, for instance, orally, topically, parenterally or byinhalation. The compounds of Formula (I) may be administered inconventional dosage forms prepared by combining a compound of Formula(I) with standard pharmaceutical carriers according to conventionalprocedures. The compounds of Formula (I) may also be administered inconventional dosages in combination with a known, secondtherapeutically; active compound. These procedures may involve mixing,granulating and compressing or dissolving the ingredients as appropriateto the desired preparation. It will be appreciated that the form andcharacter of the pharmaceutically acceptable character or diluent isdictated by the amount of active ingredient with which it is to becombined, the route of administration and other well-known variables.The carrier(s) must be “acceptable” in the sense of being compatiblewith the other ingredients of the formulation and not deleterious to therecipient thereof.

The pharmaceutical carrier employed may be, for example, either a solidor liquid. Exemplary of solid carriers are lactose, terra alba, sucrose,talc, gelatin, agar, pectin, acacia, magnesium stearate, stearic acidand the like. Exemplary of liquid carriers are syrup, peanut oil, oliveoil, water and the like. Similarly, the carrier or diluent may includetime delay material well known to the art, such as glycerylmono-stearate or glyceryl distearate alone or with a wax.

A wide variety of pharmaceutical forms can be employed. Thus, if a solidcarrier is used, the preparation can be tableted, placed in a hardgelatin capsule in powder or pellet form or in the form of a troche orlozenge. The amount of solid carrier will vary widely but preferablywill be from about 25 mg. to about 1 g. When a liquid carrier is used,the preparation will be in the form of a syrup, emulsion, soft gelatincapsule, sterile injectable liquid such as an ampule or nonaqueousliquid suspension.

Compounds of Formula (I) may be administered topically, that is bynon-systemic administration. This includes the application of a compoundof Formula (I) externally to the epidermis or the buccal cavity and theinstillation of such a compound into the ear, eye and nose, such thatthe compound does not significantly enter the blood stream. In contrast,systemic administration refers to oral, intravenous, intraperitoneal andintramuscular administration.

Formulations suitable for topical administration include liquid orsemi-liquid preparations suitable for penetration through the skin tothe site of inflammation such as liniments, lotions, creams, ointmentsor pastes, and drops suitable for administration to the eye, ear ornose. The active ingredient may comprise, for topical administration,from 0.001% to 10% w/w, for instance from 1% to 2% by weight of theformulation. It may however comprise as much as 10% w/w but preferablywill comprise less than 5% w/w, more preferably from 0.1% to 1% w/w ofthe formulation.

Lotions according to the present invention include those suitable forapplication to the skin or eye. An eye lotion may comprise a sterileaqueous solution optionally containing a bactericide and may be preparedby methods similar to those for the preparation of drops. Lotions orliniments for application to the skin may also include an agent tohasten drying and to cool the skin, such as an alcohol or acetone,and/or a moisturizer such as glycerol or an oil such as castor oil orarachis oil.

Creams, ointments or pastes according to the present invention aresemi-solid formulations of the active ingredient for externalapplication. They may be made by mixing the active ingredient infinely-divided or powdered form, alone or in solution or suspension inan aqueous or non-aqueous fluid, with the aid of suitable machinery,with a greasy or non-greasy base. The base may comprise hydrocarbonssuch as hard, soft or liquid paraffin, glycerol, beeswax, a metallicsoap; a mucilage; an oil of natural origin such as almond, corn,arachis, castor or olive oil; wool fat or its derivatives or a fattyacid such as steric or oleic acid together with an alcohol such aspropylene glycol or a macrogel. The formulation may incorporate anysuitable surface active agent such as an anionic, cationic or non-ionicsurfactant such as a sorbitan ester or a polyoxyethylene derivativethereof. Suspending agents such as natural gums, cellulose derivativesor inorganic materials such as silicaceous silicas, and otheringredients such as lanolin, may also be included.

Drops according to the present invention may comprise sterile aqueous oroily solutions or suspensions and may be prepared by dissolving theactive ingredient in a suitable aqueous solution of a bactericidaland/or fungicidal agent and/or any other suitable preservative, andpreferably including a surface active agent. The resulting solution maythen be clarified by filtration, transferred to a suitable containerwhich is then sealed and sterilized by autoclaving or maintaining at98-100° C. for half an hour. Alternatively, the solution may besterilized by filtration and transferred to the container by an aseptictechnique. Examples of bactericidal and fungicidal agents suitable forinclusion in the drops are phenylmercuric nitrate or acetate (0.002%),benzalkonium chloride (0.01%) and chlorhexidine acetate (0.01%).Suitable solvents for the preparation of an oily solution includeglycerol, diluted alcohol and propylene glycol.

Compounds of Formula (I) may be administered parenterally, that is byintravenous, intramuscular, subcutaneous intranasal, intrarectal,intravaginal or intraperitoneal administration. The subcutaneous andintramuscular forms of parenteral administration are generallypreferred. Appropriate dosage forms for such administration may beprepared by conventional techniques. Compounds of Formula (I) may alsobe administered by inhalation, that is by intranasal and oral inhalationadministration. Appropriate dosage forms for such administration, suchas an aerosol formulation or a metered dose inhaler, may be prepared byconventional techniques.

For all methods of use disclosed herein for the compounds of Formula(I), the daily oral dosage regimen will preferably be from about 0.1 toabout 80 mg/kg of total body weight, preferably from about 0.2 to 30mg/kg, more preferably from about 0.5 mg to 15 mg. The daily parenteraldosage regimen about 0.1 to about 80 mg/kg of total body weight,preferably from about 0.2 to about 30 mg/kg, and more preferably fromabout 0.5 mg to 15 mg/kg. The daily topical dosage regimen willpreferably be from 0.1 mg to 150 mg, administered one to four,preferably two or three times daily. The daily inhalation dosage regimenwill preferably be from about 0.01 mg/kg to about 1 mg/kg per day. Itwill also be recognized by one of skill in the art that the optimalquantity and spacing of individual dosages of a compound of Formula (I)or a pharmaceutically acceptable salt thereof will be determined by thenature and extent of the condition being treated, the form, route andsite of administration, and the particular patient being treated, andthat such optimums can be determined by conventional techniques. It willalso be appreciated by one of skill in the art that the optimal courseof treatment, i.e., the number of doses of a compound of Formula (I) ora pharmaceutically acceptable salt thereof given per day for a definednumber of days, can be ascertained by those skilled in the art usingconventional course of treatment determination tests.

The novel compounds of Formula (I) may also be used in association withthe veterinary treatment of mammals, other than humans, in need ofinhibition of CSBP/p38 or cytokine inhibition or production. Inparticular, CSBP/p38 mediated diseases for treatment, therapeutically orprophylactically, in animals include disease states such as those notedherein in the Methods of Treatment section, but in particular viralinfections. Examples of such viruses include, but are not limited to,lentivirus infections such as, equine infectious anaemia virus, caprinearthritis virus, visna virus, or maedi virus or retrovirus infections,such as but not limited to feline immunodeficiency virus (FIV), bovineimmunodeficiency virus, or canine immunodeficiency virus or otherretroviral infections.

The invention will now be described by reference to the followingbiological examples which are merely illustrative and are not to beconstrued as a limitation of the scope of the present invention.

Biological Examples

The cytokine-inhibiting effects of compounds of the present inventionmay be determined by the following in vitro assays:

Assays for Interleukin-1 (IL-1), Interleukin-8 (IL-8), and TumourNecrosis Factor (TNF) are well known in the art, and may be found in anumber of publications, and patents. Representative suitable assays foruse herein are described in Adams et al., U.S. Pat. No. 5,593,992, whosedisclosure is incorporated by reference in its entirety.

Interleukin-1 (IL-1)

Human peripheral blood monocytes are isolated and purified from eitherfresh blood preparations from volunteer donors, or from blood bank buffycoats, according to the procedure of Colotta et al, J Immunol, 132, 936(1984). These monocytes (1×10⁶) are plated in 24-well plates at aconcentration of 1-2 million/ml per well. The cells are allowed toadhere for 2 hours, after which time non-adherent cells are removed bygentle washing. Test compounds are then added to the cells for 1 hbefore the addition of lipopolysaccharide (50 ng/ml), and the culturesare incubated at 37° C. for an additional 24 h. At the end of thisperiod, culture supernatants are removed and clarified of cells and alldebris. Culture supernatants are then immediately assayed for IL-1biological activity, either by the method of Simon et al., J. Immunol.Methods, 84, 85, (1985) (based on ability of IL-1 to stimulate aInterleukin 2 producing cell line (IL-4) to secrete IL-2, in concertwith A23187 ionophore) or the method of Lee et al., J. ImmunoTherapy, 6(1), 1-12 (1990) (ELISA assay).

In vivo TNF Assay:

Suitable assays may be found in:

(1) Griswold et al., Drugs Under Exp. and Clinical Res.,XIX (6), 243-248(1993); or

(2) Boehm, et al., Journal Of Medicinal Chemistry 39, 3929-3937 (1996)whose disclosures are incorporated by reference herein in theirentirety.

LPS-induced TNFα Production in Mice and Rats

In order to evaluate in vivo inhibition of LPS-induced TNF(x productionin rodents, both mice and rats are injected with LPS.

Mouse Method

Male Balb/c mice from Charles River Laboratories are pretreated (30minutes) with compound or vehicle. After the 30 min. pretreat time, themice are given LPS (lipopolysaccharide from Esherichia coli Serotype055-85, Sigma Chemical Co., St Louis, Mo.) 25 ug/mouse in 25 ulphosphate buffered saline (pH 7.0) intraperitoneally. Two hours laterthe mice are killed by CO₂ inhalation and blood samples are collected byexsanguination into heparinized blood collection tubes and stored onice. The blood samples are centrifuged and the plasma collected andstored at −20° C. until assayed for TNFα by ELISA.

Rat Method

Male Lewis rats from Charles River Laboratories are pretreated atvarious times with compound or vehicle. After a determined pretreattime, the rats are given LPS (lipopolysaccharide from Esherichia coliSerotype 055-85, Sigma Chemical Co., St Louis, Mo.) 3.0 mg/kgintraperitoneally. The rats are killed by CO₂ inhalation and heparinizedwhole blood is collected from each rat by cardiac puncture 90 minutesafter the LPS injection. The blood samples are centrifuged and theplasma collected for analysis by ELISA for TNFα levels.

ELISA Method

TNFα levels were measured using a sandwich ELISA, as described inOlivera et al., Circ. Shock, 37, 301-306, (1992), whose disclosure isincorporated by reference in its entirety herein, using a hamstermonoclonal antimurine TNFα (Genzyme, Boston, Mass.) as the captureantibody and a polyclonal rabbit antimurine TNFa (Genzyme) as the secondantibody. For detection, a peroxidase-conjugated goat antirabbitantibody (Pierce, Rockford, Ill.) was added, followed by a substrate forperoxidase (1 mg/ml orthophenylenediamine with 1% urea peroxide). TNFαlevels in the plasma samples from each animal were calculated from astandard curve generated with recombinant murine TNFα (Genzyme).

LPS-Stimulated Cytokine Production in Human Whole Blood

Assay: Test compound concentrations were prepared at 10× concentrationsand LPS prepared at 1 ug/mI (final conc. of 50 ng/ml LPS) and added in50 uL volumes to 1.5 mL eppendorf tubes. Heparinized human whole bloodwas obtained from healthy volunteers and was dispensed into eppendorftubes containing compounds and LPS in 0.4 mL volumes and the tubesincubated at 37 C. Following a 4 hour incubation, the tubes werecentrifuged at 5000 rpm for 5 minutes in a TOMY microfuge, plasma waswithdrawn and frozen at −80 C.

Cytokine measurement: IL-1 and/or TNF were quantified using astandardized ELISA technology. An in-house ELISA kit was used to detecthuman IL-1 and TNF. Concentrations of IL-1 or TNF were determined fromstandard curves of the appropriate cytokine and IC50 values for testcompound (concentration that inhibited 50% of LPS-stimulated cytokineproduction) were calculated by linear regression analysis.

CSBP/p38 Kinase Assay:

This assay measures the CSBP/p38-catalyzed transfer of ³²p from[a-³²P]ATP to threonine residue in an epidermal growth factor receptor(EGFR)-derived peptide (T669) with the following sequence:KRELVEPLTPSGEAPNQALLR (residues 661-681). (See Gallagher et al.,“Regulation of Stress Induced Cytokine Production by PyridinylImidazoles: Inhibition of CSBP Kinase”, BioOrganic & MedicinalChemistry, 1997, 5, 49-64).

Reactions were carried in round bottom 96 well plate (from Coming) in a30 ml volume. Reactions contained (in final concentration): 25 mM Hepes,pH 7.5; 8 mM MgCl₂; 0.17 mM ATP (the Km_([ATP]) of p38 (see Lee et al.,Nature 300, n72 pg. 639-746 (Dec. 1994)); 2.5 uCi of [g-32P]ATP; 0.2 mMsodium orthovanadate; 1 mM DTT; 0.1% BSA; 10% glycerol; 0.67 mM T669peptide; and 2-4 nM of yeast-expressed, activated and purified p38.Reactions were initiated by the addition of [gamma-32P]Mg/ATP, andincubated for 25 min. at 37° C. Inhibitors (dissolved in DMSO) wereincubated with the reaction mixture on ice for 30 minutes prior toadding the 32P-ATP. Final DMSO concentration was 0.16%. Reactions wereterminated by adding 10 ul of 0.3 M phosphoric acid, and phosphorylatedpeptide was isolated from the reactions by capturing it on p81phosphocellulose filters. Filters were washed with 75 mM phosphoricacids, and incorporated 32P was quantified using beta scintillationcounter. Under these conditions, the specific activity of p38 was400-450 pmol/pmol enzyme, and the activity was linear for up to 2 hoursof incubation. The kinase activity values were obtained aftersubtracting values generated in the absence of substrate which were10-15% of total values.

Representative compounds of Formula (I), Examples 1 to 14 alldemonstrated positive inhibitory activity of an IC₅₀ of <50 uM in thiskinase assay or a similar binding assay.

Prostaglandin Endoperoxide Synthase-2 (PGHS-2) Assay:

This assay describes a method for determining the inhibitory effects ofcompounds of Formula (I) on human PGHS-2 protein expression in LPSstimulated human monocytes. A suitable assay for PGHS-2 proteinexpression may be found in a number of publications, including U.S. Pat.No. 5,593,992 whose disclosure is incorporated herein by reference.

TNF-α in Traumatic Brain Injury Assay

This assay provides for examination of the expression of tumor necrosisfactor mRNA in specific brain regions which follow experimentallyinduced lateral fluid-percussion traumatic brain injury (TBI) in rats.Since TNF-α is able to induce nerve growth factor (NGF) and stimulatethe release of other cytokines from activated astrocytes, thispost-traumatic alteration in gene expression of TNF-α plays an importantrole in both the acute and regenerative response to CNS trauma. Asuitable assay may be found in WO 97/35856 whose disclosure isincorporated herein by reference.

CNS Injury Model for IL-b mRNA

This assay characterizes the regional expression of interleukin-1β(IL-1β) mRNA in specific brain regions following experimental lateralfluid-percussion traumatic brain injury (TBI) in rats. Results fromthese assays indicate that following TBI, the temporal expression ofIL-1β mRNA is regionally stimulated in specific brain regions. Theseregional changes in cytokines, such as IL-1β play a role in thepost-traumatic pathologic or regenerative sequelae of brain injury. Asuitable assay may be found in WO 97/35856 whose disclosure isincorporated herein by reference.

Angiogenesis Assay:

Described in WO 97/32583, whose disclosure is incorporated herein byreference, is an assay for determination of inflammatory angiogenesiswhich may be used to show that cytokine inhibition will stop the tissuedestruction of excessive or inappropriate proliferation of bloodvessels.

Rhinovirus Methods:

Cell lines, rhinovirus serotype 39, and influenza virus A/PR/8/34 arepurchased from American Type Culture Collection (ATCC). BEAS-2B cellswere cultured according to instructions provided by ATCC using BEGM(bronchial epithelial growth media) purchased from Clonetics Corp. HELAcell cultures, used for detection and titration of virus, are maintainedin Eagle's minimum essential media containing 10% fetal calf serum, 2 mM1-glutamine, and 10 mM HEPES buffer (MEM).

A modification of the method reported by Subauste et al., Supra, for invitro infection of human bronchial epithelial cells with rhinovirus isused in these studies. BEAS-2B cells (2×10⁵/well) are cultured incollagen-coated wells for 24 hours prior to infection with rhinovirus.Rhinovirus serotype 39 is added to cell cultures for one hour incubationat 34° C. after which inoculum is replaced with fresh media and culturesare incubated for an additional 72 hours at 34° C. Supernatants arecollected at 72 hours post-infection and are assayed for cytokineprotein concentration by ELISA using commercially available kits (R&DSystems). Virus yield is also determined from culture supernatants usinga microtitration assay in HELA cell cultures (Subauste et al., supra1995). In cultures treated with p38 kinase inhibitors, drug is added 30minutes prior to infection. Stocks of compounds are prepared in DMSO (10mM drug) and stored at −20° C.

For detection of p38 kinase, cultures are incubated in basal mediawithout growth factors and additives to reduce endogenous levels ofactivated p38 kinase. Cells are harvested at various timepoints afteraddition of rhinovirus. Detection of tyrosine phosphorylated p38 kinaseby immunoblot is analyzed by a commercially available kit and isperformed according to the manufacturer's instructions (PhosphoPlus p38MAPK Antibody Kit: New England BioLabs Inc.).

In some experiments, BEAS-2B cells may be infected with influenza virus(strain A/PR/8/34) in place of rhinovirus. Culture supernatant isharvested 48 and 72 hour post-infection and tested by ELISA for cytokineas described above.

Cells and Virus: Influenza A/PR/8/34 sub type H1N1 (VR-95 American TypeCulture Collection, Rockville, Md.) is grown in the allantoic cavity of10 day old chicken eggs. Following incubation at 37° C., andrefrigeration for 2½ hours at 4° C., allantoic fluid is harvested,pooled, and centrifuged (1,000 rcf; 15 min; 4° C.) to remove cells.Supernatent is aliquoted and stored at −70° C. The titer of the stockculture of virus is 1.0×10¹⁰ Tissue Culture Infective Dose/ml (TCID₅₀)

Inoculation procedure: Four-six week old female Balb/cAnNcrlBr mice areobtained from Charles River, Raleigh, N.C. Animals are infectedintranasally. Mice are anesthetized by intraperitioneal injection ofKetamine (40 mg/kg; Fort Dodge Labs, Fort Dodge, Iowa) and Xylazine (5mg/kg; Miles, Shawnee Mission, Kans.) and then inoculated with 100TCID50 of PR₈ diluted in PBS in 20 ul. Animals are observed daily forsigns of infection.

Virus titration: At various times post infection, animals are sacrificedand lungs are aseptically harvested. Tissues are homogenized, in vialscontaining 1 micron glass beads (Biospec Products, Bartlesville, Okla.)and 1 ml. of Eagles minimal essential medium. Cell debris is cleared bycentrifugation at 1,000 rcf for 15 minutes at 4° C., and supernatantsare serially diluted on Madin-Darby canine kidney (MDCK) cells. After 5days of incubation at 37° C. (5% CO₂), 50 μl of 0.5% chick red bloodcells are added per well, and agglutination is read after 1 hour at roomtemperature. The virus titer is expressed as 50% tissue cultureinfective dose (TCID₅₀) calculated by logistic regression.

ELISA: Cytokine levels are measured by quantitative ELISA usingcommercially available kits. Ear samples are homogenized using a tissueminser in PBS. Cell debris is cleared by centrifugation at 14,000 rpmfor 5 minutes. The cytokine concentrations and thresholds are determinedas described by the manufacturer; IL-6, IFN-γ, and KC (R&D Systems,Minneapolis, Minn.).

Myeloperoxidase Assay: Myeloperoxidase (MPO) activity is determinedkinetically as described by Bradley et al. (1982). Briefly, rabbitcornea are homogenized in Hexadecyl Trimethyl-Ammonium Bromide (HTAB)(Sigma Chemical Co. St. Louis, Mo.) which is dissolved in 0.5 mPotassium phosphate buffer (J. T. Baker Scientific, Phillipsburg, N.J.).Following homogenization, the samples are subjected tofreeze-thaw-sonication (Cole-Parmer 8853, Cole-Parmer, Vernon Hills,Ill.) 3 times. Suspensions are then cleared by centrifugation at12,500×g for 15 minutes at 4° C. MPO enzymatic activity is determined bycolormetric change in absorbance during a reaction of O-Dianisidinedihydrochloride (ODI) 0.175 mg/ml (Sigma Chemical Co. St. Louis, Mo)with .0002% Hydrogen peroxide (Sigma Chemical Co. St. Louis, Mo.).Measurements are performed by using a Beckman Du 640 Spectrophotometer(Fullerton, Calif.) fitted with a temperature control device. 50 ul ofmaterial to be assayed is added to 950 ul of ODI and change inabsorbance is measured at a wave length of 460 nm for 2 minutes at 25°C.

Whole Body Plethysomography: Influenza virus infected mice are placedinto a whole body plethysomograph box with an internal volume ofapproximately 350-ml. A bias airflow of one 1/min is applied to the boxand flow changes were measured and recorded with a Buxco XA dataacquisition and respiratory analysis system (Buxco Electronics, Sharon,Conn.). Animals are allowed to acclimate to the plethysmograph box for 2min. before airflow data is recorded. Airway measurements are calculatedas Penh (enhanced pause). Penh has previously been shown as an index ofairway obstruction and correlates with increased intrapleural pressure.The algorithm for Penh calculation is as follows: Penh=[(expiratorytime/relaxation time)-1]×(peak expiratory flow/peak inspiratory flow)where relaxation time is the amount of time required for 70% of thetidal volume to be expired.

Determination of arterial oxygen saturation. A Nonin veterinary handheld pulse oximeter 8500V with lingual sensor (Nonin Medical, Inc.,Plymouth Minn.) is used to determine daily arterial oxygen saturation %SpO2 as described (Sidwell et al. 1992 Antimicrobial Agents andChemotherapy 36:473-476).

Results:

Inhibition of cytokine production by specific inhibitors of p38 MAPkinase:

Consistent with published reports, IL-6, IL-8, and GM-CSF are detected72 hours post-infection of BEAS-2B cells with rhinovirus-39(multiplicity of infection; MOI 1.0) (FIG. 1). Production of IL-6, IL-8,and GM-CSF is not mediated through IL-1 or TNF produced in response torhinovirus infection since addition of neutralizing antibodies to IL-1and TNF to the infected cultures did not reduce the amount of IL-6, IL-8or GM-CSF produced (not shown). Productive infection of cells isconfirmed by titering infectious supernatants from BEAS-2B cells on HELAmonolayers. There is low but consistent replication of virus during the72 hour culture period resulting in 1.22±0.3 log₁₀ TCID₅₀ increase overthe initial input inoculum (n=6 experiments).

To investigate the role of p38 kinase signal transduction inrhinovirus-induced cytokine production by epithelial cells, specific p38kinase inhibitors of Formula (I) may be tested for their ability toinhibit cytokine production in the rhinovirus-infected BEAS-2B cellcultures.

Further data on this method may be found in PCT application U.S. Ser.No. 00/125386, filed Sep. 15, 2000 whose disclosure is incorporatedherein by reference in its entirety.

Activation of p38 kinase by rhinovirus infection:

The presence of tyrosine phosphorylated p38 kinase is measured byinmmunoblot at various times after the addition of virus to BEAS-2Bcultures. Rhinovirus infection of BEAS-2B cells results in an increasein phosphorylated p38 kinase that was both dose and time-dependent.Increases in phosphorylated p38 kinase is evident by 15 minutes postexposure to rhinovirus-39 (MOI 10), appeared to peak by 30 minutes afteraddition of virus and remained elevated 60 minutes post-infection (FIG.3). In addition, rhinovirus-induced tyrosine phosphorylation of p38kinase was dose-dependent. When cells are cultured in the absence ofvirus, there was no increase in the amount of tyrosine phosphorylationof p38 kinase at any of the timepoints tested. Overall levels of p38kinase protein were comparable between all the groups indicating thatvirus infection caused phosphorylation of p38 kinase without de novosynthesis of protein.

Effects on in vitro influenza virus infection:

Exposure of BEAS-2B cells with influenza virus (A/PR/8/34; MOI 1.0) alsoresults in elaboration of IL-8 and IL-6 as measured 48-72 hourspost-infection, although the secreted protein levels are lower than thatobtained with rhinovirus infection.

Cigarette Smoke Exposure Model

A murine model of cigarette smoke inhalation was developed to explore arelationship to leukocyte trafficking and lung chemokine and cytokineproduction. Balb/c mice are exposed to smoke generated from commercialunfiltered cigarettes for a specified period of time and samples areobtained at varying times during the post-exposure. This model isdemonstrated in greater detail as shown below, in contrast to othersmoke extract models known in the art.

A model of cigarette smoke exposure in the mouse is established in whichmice are placed 6 at a time into a small animal plexiglass dosingchamber attached to a peristaltic pump whose intake is connected to aholder for a commercial unfiltered cigarette (Lucky Strike™). Along withfresh air, smoke is delivered into the chamber until the cigarette isconsumed (approximately 5 minutes). Varying numbers of cigarettes (2-4per day, 2-3 hr apart) are utilized for 1-3 consecutive days. Animalsare euthanized by pentobarbital overdose approximately 18 hours afterthe final exposure. Bronchoalveolar lavage with phosphate-bufferedsaline is performed for inflammatory cell enumeration, and BAL aliquotsand lungs .are frozen for cytokine analysis. Smoke exposure results intime- and cigarette number-related increases in airway neutrophils, andlung chemokine (KC) and cytokine (IL-6) content.

To evaluate the role of a p38 MAP kinase inhibitor in this inflammatoryresponse, mice are treated with a p38 kinase inhibitor, a compound ofFormula (I) at approximatetly a 30 mg/kg, p.o. b.i.d. Reduction in lungKC (a murine homolog of IL-8) levels are assesed 1 day after exposure(prior to neutrophilia), and attenuated airway neutrophilia and lungIL-6 levels are assessed following 3 days of cigarette exposure.

Hypertussive Cough Models

Described below is an example of how to determine the usefulness of p38inhibitors in the treatment of hypertussive disorders or inflammationenhanced cough.

The directed antitussive activity of the compound in question if firstassessed, by a 10 to 30 minute pretreatment period by intraperitonealinjection or a 1 hour pretreatment period for oral administration. Theanimals (guinea pigs) are then subjected to an inhaled citricacid-induced cough challenge. The Citric Acid Induced Cough Model isshown in FIG. 2.

The effects of the compound are then assessed on the hypertussiveresponse that occurs 72 hours post aerosol exposure to antigen or LTD4exposure. Treatment of the animals occurs with the drug prior and/orafter antigen or LTD4 challenge, but not on the day of citric acidchallenge. The antigen or LTD4 induced hypertussive model is shown inFIG. 3.

The effects of known antitussive agents, dextromethorphan and codeine onCitric Acid Induced Cough in Guinea Pigs is shown in FIG. 4.

Inhalation of citric acid (CA; 0.4% for 1 minute) induces 11 to 15coughs during the exposure and 12-minute monitoring period in consciousguinea pigs. Exposure of sensitized animals to inhaled ovalbuminresulted in a hypertussive state (50-80% increae in CA-induced coughincidence) for several days, which positively correlated with airwayesoinophilia determined by bronchoalveolar lavage.

Similarly, inhalation of LTD4 (10 ug/ml for 1 minute) increases coughincidence and airway esoinophils 72 hours after exposure.

SYNTHETIC EXAMPLES

The invention will now be described by reference to the followingexamples which are merely illustrative and are not to be construed as alimitation of the scope of the present invention. All temperatures aregiven in degrees centigrade, all solvents are highest available purityand all reactions run under anhydrous conditions in an argon atmosphereunless otherwise indicated.

In the Examples, all temperatures are in degrees Centigrade (° C.). Massspectra were performed upon a VG Zab mass spectrometer using fast atombombardment or on a micromass platform electrospray ionization massspectrometer in the positive ion mode using 95:5 CH₃CN/CH₃OH with 1%formic acid as the carrier solvent, unless otherwise indicated. ¹H-NMR(hereinafter “NMR”) spectra were recorded at 250 MHz using a Bruker AM250 or Am 400 spectrometer. Multiplicities indicated are: s=singlet,d=doublet, t=triplet, q=quartet, m=multiplet and br indicates a broadsignal. Sat. indicates a saturated solution, eq indicates the proportionof a molar equivalent of reagent relative to the principal reactant.

Flash chromatography is run over Merck Silica gel 60 (23-400 mesh).

Example 1 1-Cyclohexyl-5-phenyl-3.4-dihydro-(1H)-guinazolin-2-one

a) 2-Fluoro-6-phenylbenzonitrile

The compound, 2-Bromo-6-fluorobenzonitrile, the procedure for thepreparation of which may be found in Hynes, John B. et. al., J.Hetercycl. Chem. 1988, 25, 1173-7, (2.0 g, 10.0 mmol) and phenylboronicacid (3.66 g, 30 mmol) were added to a mixture of toluene (80 mL),methanol (20 mL), and 2 M aqueous sodium carbonate (20 mL). This mixturewas refluxed while stirring under argon for 15 min. The resultingmixture was cooled to room temperature andtetrakis(triphenylphosphine)palladium(0) (1.16 g, 1 mmol) was added. Theresulting mixture was then refluxed for 12 hours while stirring underargon. The solvent was removed in vacuo and the residue was partitionedbetween ethyl acetate and water. The organic phase was washed with water(2×), brine (1×), dried over anhydrous Na₂SO₄, filtered, and evaporatedto give the crude product which was then chromatographed on silica geleluted with 0-10% methylene chloride in hexane. Recrystallization frommethylene chloride hexane gave the title product as a white crystallinesolid. mp 78-79° C.

b) 2-Cyclohexylamino-6-phenylbenzonitrile

The product of example la above, 2-fluoro-6-phenylbenzonitrile, (0.1 g,0.51 mmol), and cyclohexylamine (1 mL) were combined in a sealed tubethat had been flushed with argon. The tube was heated in an oil bath to100° C. for 1.6 hours. The excess cyclohexylamine was evaporated, andthe residue was flash chromatographed on silica gel eluted with 0-2%ethyl acetate in hexane, and then recrystallized from methylenechloride/hexane to give the title product as a white crystallineproduct. mp 125-126° C.

c) 2-Cyclohexylamino-6-(phenyl)benzylamine

The product of example 1b above, 2-cyclohexylamino-6-phenylbenzonitrile,(0.095 g, 0.34 mmol) was dissolved in dry tetrahydrofuran (20 mL) andstirred under argon at room temperature. Lithium aluminum hydride (0.136g, 3.4 mmol) was added and the mixture heated to reflux under argon for9 hours. The reaction mixture was cooled to room temperature andanhydrous sodium sulfate was added followed by the addition of a freshlyprepared saturated solution of anhydrous sodium sulfate. The solvent wasevaporated and the residue triturated with ethyl acetate, filtered andevaporated. Flash chromatography on silica gel eluted with 0-2% methanolin methylene chloride followed by recrystallization from ethylacetate/hexane gave the title compound as a white crystalline solid. mp95-96° C.

d) 1-Cyclohexyl-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

The product of example 1c above,2-cyclohexylamino-6-(phenyl)benzylamine, (0.060 g, 0.21 mmol) wasdissolved in methylene chloride (10 mL) and cooled in an ice bath whilestirring under argon. Phosgene, 20% in toluene, (0.13 mL, 0.25 mmol) wasadded followed by triethylamine (0.7 mL). The ice bath was removed andthe mixture stirred for 30 minutes at room temperature. The solventswere evaporated, and the residue was partitioned between ethyl acetateand water. The organic phase was washed with brine, dried over anhydrousNa₂SO₄ filtered, and evaporated to give the crude product which wasrecrystallized from methylene chloride/hexane to give the title compoundas a white crystalline compound. mp 229-231° C.

Example 2 1-Isopropyl-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one

a) 2-Isopropylamino-6-phenylbenzonitrile

The product of example 1a above, 2-fluoro-6-phenylbenzonitrile, (0.12 g,0.61 mmol), and isopropylamine (3 mL) were combined in a sealed tubethat had been flushed with argon. The tube was heated in an oil bath to95° C. for 7 days. The excess isopropylamine was evaporated, .and theresidue was flash chromatographed on silica gel eluted with 0-3% ethylacetate in hexane, and then crystallized from hexane to give the titleproduct as a white crystalline product. ES (+) MS m/e=237 (MH+)

b) 2-Isopropylamino-6-(phenyl)benzylamine

The product of example 2a above, 2-isopropylamino-6-phenylbenzonitrile,(0.095 g, 0.4 mmol) was dissolved in dry tetrahydrofuran (20 mL) andstirred under argon at room temperature. Lithium aluminum hydride (0.160g, 4 mmol) was added and the mixture heated to reflux under argon for 16hours. The reaction mixture was cooled to room temperature and anhydroussodium sulfate was added followed by the addition of a freshly preparedsaturated solution of anhydrous sodium sulfate. The solvent wasevaporated and the residue triturated with ethyl acetate, filtered andevaporated. Flash chromatography on silica gel eluted with 0-3% methanolin methylene chloride followed by recrystallization from ethylacetate/hexane gave the title compound as a white crystalline solid. ES(+) MS m/e=241 (MH+)

c) 1-Isopropyl-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

The product of example 2b above, 2-isopropylamino-6-(phenyl)benzylamine,(0.090 g, 0.37 mmol) was dissolved in methylene chloride (10 mL) andcooled in an ice bath while stirring under argon. Phosgene, 20% intoluene, (0.235 mL, 0.45 mmol) was added followed by triethylamine (1.0mL). The ice bath was removed and the mixture stirred for 30 minutes atroom temperature. The solvents were evaporated, and the residue waspartitioned between ethyl acetate and water. The organic phase waswashed with brine, dried over anhydrous Na₂SO₄ filtered, and evaporatedto give the crude product which was recrystallized from methylenechloride/hexane to give the title compound as a white crystallinecompound. mp 182-183° C.

Example 3(+/−)-1-[1-(Phenyl)ethyl]-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

a) (+/−)-2-[(1-phenyl)ethylamino]-6-phenylbenzonitrile

The product of example 1a above, 2-fluoro-6-phenylbenzonitrile, (0.118g, 0.6 mmol), and (+/−)-1-(phenyl)ethylamine (0.5 mL) were combined in asealed tube that had been flushed with argon. The tube was heated in anoil bath to 120° C. for 18 hours. The reaction was not complete at thistime, so the reaction was heated at 150° C. for an additional 48 hours.Flash chromatography on silica gel eluted with 0-3% ethyl acetate inhexane gave the title product. ES (+) MS m/e=299 (MH+)

b) (+/−)-2-[(1-phenyl)ethylamino]-6-(phenyl)benzylamine

The product of example 3a above,(+/−)-2-[(1-phenyl)ethylamino]-6-phenybenzonitrile, (0.13 g, 0.43 mmol)was dissolved in dry tetrahydrofuran (10 mL) and stirred under argon atroom temperature. Lithium aluminum hydride (0.172 g, 4.3 mmol) was addedand the mixture heated to reflux under argon for 20 hours. The reactionmixture was cooled to room temperature and anhydrous sodium sulfate wasadded followed by the addition of a freshly prepared saturated solutionof anhydrous sodium sulfate. The solvent was evaporated and the residuetriturated with ethyl acetate, filtered and evaporated. Flashchromatography on silica gel eluted with 0-3% methanol in methylenechloride followed by recrystallization from ethyl acetate/hexane gavethe title compound as a white crystalline solid. ES (+) MS n/e=303 (MH+)

c) (+/−)-1-[1-(Phenyl)ethyl]-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

The product of example 3b above,(+/−)-2-[1-(Phenyl)ethylamino-6-(phenyl)benzylamine, (0.121 g, 0.4 mmol)was dissolved in methylene chloride (10 mL) and cooled in an ice bathwhile stirring under argon. Phosgene, 20% in toluene, (0.25 mL, 0.48mmol) was added followed by triethylamine (1.0 mL). The ice bath wasremoved and the mixture stirred for 30 minutes at room temperature. Thesolvents were evaporated, and the residue was partitioned between ethylacetate and water. The organic phase was washed with brine, dried overanhydrous Na₂SO₄ filtered, and evaporated to give the crude productwhich was recrystallized from methylene chloride/hexane to give thetitle compound as a white crystalline solid. mp 169-171° C.

Example 4(R)-1-[1-(Phenyl)ethyl]-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

Following the procedures of example 3 except for substituting(R)-1-(phenyl)ethylamine for (+/−)-1-(phenyl)ethylamine in step 3a gavethe title compound as a white crystalline solid. mp 131-134° C.

Example 5(S)-1-[1-(Phenyl)ethyl]-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

Following the procedures of example 3 except for substituting(S)-1-(phenyl)ethylamine for (+/−)-1-(phenylethylamine in step 3a gavethe title compound as a white crystalline solid. mp 136-138° C.

Example 6 1-(2-Phenethyl)-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

Following the procedures of example 3 except for substituting2-phenylethylamine for (+/−)-1-(phenyl)ethylamine in step 3a gave thetitle compound as a white crystalline solid. mp 152-153° C.

Example 7 1-Benzyl-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

a) 2-Benzylamino-6-phenylbenzonitrile

The product of example 1a above, 2-fluoro-6-phenylbenzonitrile, (0.118g, 0.6 mmol), and benzylamine (0.5 mL) were combined in a sealed tubethat had been flushed with argon. The tube was heated in an oil bath to150° C. for 24 hours. The residue was dissolved in methylene chloride(10 mL) and treated with an excess of polystyrene supported isocyanateresin for 30 minutes to remove the excess benzylamine. Filtration andevaporation gave the title product. ES (+) MS m/e=285 (MH+)

b) 2-Benzylamino-6-(phenyl)benzylamine

The product of example 7a above, 2-benzylamino-6-phenylbenzonitrile, wasdissolved in dry tetrahydrofuran (10 mL) and stirred under argon at roomtemperature. Lithium aluminum hydride (0.175 g, 4.3 mmol) was added andthe mixture heated to reflux under argon for 20 hours. The reactionmixture was cooled to room temperature and anhydrous sodium sulfate wasadded followed by the addition of a freshly prepared saturated solutionof anhydrous sodium sulfate. The solvent was evaporated and the residuetriturated with ethyl acetate, filtered and evaporated to give the titlecompound. ES (+) MS m/e=272(M-NH2+)

c) 1-Benzyl-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

The product of example 7b above, 2-benzyl-6-(phenyl)benzylamine,(assumed 0.4 mmol) was dissolved in methylene chloride (10 mL) andcooled in an ice bath while stirring under argon. Phosgene, 20% intoluene, (0.376 mL, 0.72 mmol) was added followed by triethylamine (2.0mL). The ice bath was removed and the mixture stirred for 30 minutes atroom temperature. The solvents were evaporated, and the residue waspartitioned between ethyl acetate and water. The organic phase waswashed with brine, dried over anhydrous Na₂SO₄ filtered, and evaporatedto give the crude product which was flash chromatographed on silica geleluted with 0-2% methanol in methylene chloride, and then recrystallizedfrom methylene chloride/hexane to give the title compound as a whitecrystalline solid. mp 198-199° C.

Example 81-(2-Morpholin-4yl-ethyl-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

The product of example 1a above, 2-fluoro-6-phenylbenizonitrile, (0.118g, 0.6 mmol), and 4-(2-aminoethyl)morpholine (0.5 mL) were combined in asealed tube that had been flushed with argon. The tube was heated in anoil bath to 150° C. for 24 hours. Most of the excess amine was removedin vacuo. The residue was taken up in tetrahydrofuran (10 mL), treatedwith lithium aluminum hydride (0.175 g, 4.3 mmol), and the mixtureheated to reflux under argon for 20 hours. The reaction mixture wascooled to room temperature and anhydrous sodium sulfate was addedfollowed by the addition of a freshly prepared saturated solution ofanhydrous sodium sulfate. The solvent was evaporated and the residuetriturated with ethyl acetate, filtered and evaporated. The residue wastaken up in methylene chloride (5 mL) and cooled in an ice bath whilestirring under argon. Phosgene, 20% in toluene, (0.375 mL, 0.72 mmol)was added followed by triethylamine (2.0 mL). The ice bath was removedand the mixture stirred for 30 minutes at room temperature. The solventswere evaporated, and the residue was partitioned between ethyl acetateand water. The organic phase was washed with brine, dried over anhydrousNa₂SO₄ filtered, and evaporated to give the crude product which wasflash chromatographed on silica gel eluted with 0-2% methanol inmethylene chloride, and then recrystallized from methylenechloride/hexane to give the title product. mp 228-229° C.

Example 91-[2-(2-Pyridinyl)ethyl]-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-oneTrifluoroacetate

Following the procedure of example 8 except for substituting2-(2-pyridinyl)ethylamine for 4-(2-aminoethyl)morpholine andpurification by reverse phase hplc purification using anacetonitrile/water/+0.1%trifluoroacetic acid gradient instead of flashchromatography on silica gel gave the title compound. ES (+) MS m/e=330(MH+)

Example 101-(2-Morpholin4-yl-propyl)-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-oneTrifluoroacetate

Following the procedure of example 8 except for substituting4-(3-aminopropyl)morpholine for 4-(2-aminoethyl)morpholine andpurification by reverse phase hplc purification using anacetonitrile/water/+0.1%trifluoroacetic acid gradient instead of flashchromatography on silica gel gave the title compound. ES (+) MS m/e=352(MH+)

Example 111-(1-Methylpyrrol-2-ylethyl)-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

Following the procedure of example 8 except for substituting1-methylpyrrol-2-ylethylamine for 2-morpholin-4-yl-ethylamine gave thetitle compound. mp 158-161° C.

Example 12 1,5-Diphenyl-3,4-dihydro-(1H)-quinazolin-2-one

a) 2-Fluoro-6-phenylbenzonitrile

The compound, 2-Bromo-6-fluorobenzonitrile, the procedure for thepreparation of which may be found in Hynes, John B. et. al., J.Hetercycl. Chem. 1988, 25, 1173-7, (2.0 g, 10.0 mmol) and phenylboronicacid (3.66 g, 30 mmol) were added to a mixture of toluene (80 mL),methanol (20 mL), and 2 M aqueous sodium carbonate (20 mL). This mixturewas refluxed while stirring under argon for 15 min. The resultingmixture was cooled to room temperature andtetrakis(triphenylphosphine)palladium(0) (1.16 g, 1 mmol) was added.The. resulting mixture was then refluxed for 12 hours while stirringunder argon. The solvent was removed in vacuo and the residue waspartitioned between ethyl acetate and water. The organic phase waswashed with water (2×), brine (1×), dried over anhydrous Na₂SO₄,filtered, and evaporated to give the crude product which was thenchromatographed on silica gel eluted with 0-10% methylene chloride inhexane. Recrystallization from methylene chloride hexane gave the titleproduct as a white crystalline solid. mp 78-79° C.

b) 2-Phenylamino-6-phenylbenzonitrile

Aniline (0.168 g, 1.8 mmol) was dissolved in dimethyl sulfoxide (3 mL)and stirred under argon at room temperature. Sodium hydride (0.0043 g,1.8 mmol) was added and a deep purple color developed while the mixturewas stirred at room temperature for 1 hour. The mixture was cooled to 0°C. in an ice bath and the product of example 12a above,2-fluoro-6-phenylbenzonitrile, (0.295 g, 1.5 mmol) dissolved in dimethylsulfoxide (2 mL) was added and the mixture stirred at room temperatureovernight. The solvent was evaporated in vacuo, and the residue waspartitioned between ethyl acetate and water. The organic phase waswashed with water (5×), brine (1×) dried over anhydrous Na₂SO₄ filtered,and evaporated to give the crude product which was then chromatographedon silica gel eluted with 0-30% CH₂Cl₂ in hexane to give the titlecompound as a white amorphous solid. ES (+) MS m/e=271 (MH+)

c) 2-Phenyl-6-(phenylamino)benzylamine

The product of example 1b above, 2-phenylamino-6-phenylbenzonitrile,(0.13 g, 0.48 mmol) was dissolved in dry tetrahydrofuran (20 mL) andstirred under argon in a room temperature water bath. Lithium aluminumhydride (0.182 g, 4.8 mmol) was added rapidly. After 5 minutes at roomtemperature the reaction mixture was heated to 60° C. for 2 hours. Thereaction mixture was cooled to room temperature and anhydrous sodiumsulfate was added followed by the addition of a freshly preparedsaturated solution of anhydrous sodium sulfate. The solvent wasevaporated and the residue triturated with ethyl acetate, filtered andevaporated. Flash chromatography on silica gel eluted with 0-2% methanolin methylene chloride gave the title compound as a white amorphoussolid. mp 115-116° C.

d) 1.5-Diphenyl-3,4-dihydro-(1H)-guinazolin-2-one

The product of example 12c above, 2-Phenyl-6-(phenylamino)benzylamine,(0.095 g, 0.35 mmol)was dissolved in dry tetrahydrofuran (3 mL) andstirred under argon at room temperature. 1,1′-Carbonyldiimidazole (0.073g, 0.45 mmol) dissolved in dry tetrahydrofuran (2 mL) was added and themixture stirred at room temperature overnight. The solvent wasevaporated in vacuo, to give the crude product which was flashchromatographed on silica gel eluted with 0-2% methanol in methylenechloride to give to the title compound as a white amorphous solid. mp234-235° C.

Example 131-(2,6-Difluorphenyl)-5-phenyl-3,4-dihydro-(1H)-guinazolin-2-one

a) 2-[(2,6-Difluorophenyl)amino]-6-phenylbenzonitrile

Following the procedure of example 12b above except for using2,6-difluoroaniline in place of aniline gave the title compound as awhite crystalline solid. mp 139-141° C.

b) 1-(2,6-Difluorphenyl)-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one

Following the procedure of example 1c and 1d above except for using2-[(2,6-Difluorophenyl)amino]-6-phenylbenzonitrile in place of2-cyclohexylamino-6-phenylbenzonitrile in step 1c gave the titlecompound as a white crystalline solid. mp 247-249° C.

Example 14 1-Phenyl-5-(2-methylphenyl)-3,4-dihydro-(1H)-quinazolin-2-onemp 236-238° C.

a) 2-fluoro-6-(2-methylphenyl)benzonitrile

Following the procedure of example 12a except substituting2-methylphenylboronic acid for phenylboronic acid gave the titlecompound. mp 57-58° C.

b) 2-Phenylamino-6-(2-methylphenyl)benzonitrile

Following the procedure of example 12b except substituting2-fluoro-6-(2-methylphenyl)benzonitrile for2-fluoro-6-phenylbenzonitrile gave the title compound. mp 125-126° C.

c) 2-Phenyl-6-(2-methylphenylamino)benzylamine

Following the procedure of example 12c except substituting2-phenylamino-6-(2-methylphenyl)benzonitrile for2-phenylamino-6-phenylbenzonitrile gave the title compound. mp 103-105°C.

d) 1-Phenyl-5-(2-methylphenyl)-3,4-dihydro-(1H)-guinazolin-2-one

Following the procedure of example 12d except substituting2-phenyl-6-(2-methylphenylamino)benzylamine for2-phenyl-6-(phenylamino)benzylamine gave the title compound. mp 236-238°C.

All publications, including but not limited to patents and patentapplications, cited in this specification are herein incorporated byreference as if each individual publication were specifically andindividually indicated to be incorporated by reference herein as thoughfully set forth.

The above description fully discloses the invention including preferredembodiments thereof. Modifications and improvements of the embodimentsspecifically disclosed herein are within the scope of the followingclaims. Without further elaboration, it is believed that one skilled inthe are can, using the preceding description, utilize the presentinvention to its fullest extent. Therefore, the Examples herein are tobe construed as merely illustrative and not a limitation of the scope ofthe present invention in any way. The embodiments of the invention inwhich an exclusive property or privilege is claimed are defined asfollows.

What is claimed is:
 1. A compound of the formula:

wherein R₁ is a phenyl, naphth-1-yl, naphth-2-yl, or heteroaryl ring,which ring is optionally substituted independently by one or moresubstituents selected from halogen, C₁₋₄ alkyl, halo-substituted-C₁₋₄alkyl, cyano, nitro, (CR₁₀R₂₀)_(v)NR₄R₁₄, (CR₁₀R₂₀)_(v)C(Z)NR₄R₁₄,(CR₁₀R₂₀)_(v)C(Z)OR₈, (CR₁₀R₂₀)_(v)COR₃, (CR₁₀R₂₀)_(v)C(O)H, SR₅,S(O)R₅, S(O)₂R₅, (CR₁₀R₂₀)_(v)OR₈, ZC(Z)R₁₁, NR₁₀C(Z)R₁₁, orNR₁₀S(O)₂R₇; R₂ is a C₁₋₁₀ alkyl, C₃₋₇ cycloalkyl, C₃₋₇cycloalkylC₁₋₁₀alkyl, C₅₋₇ cycloalkenyl, C₅₋₇ cycloalkenyl C₁₋₁₀ alkyl, arylC₁₋₁₀alkyl, heteroarylC₁₋₁₀-alkyl, or heterocyclylC₁₋₁₀ alkyl moiety, whichmoiety is optionally substituted one or more times independently withC₁₋₁₀ alkyl, halo-substituted C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl,C₃₋₇ cycloalkyl, C₃₋₇cycloalkylC₁₋₁₀ alkyl, C₅₋₇ cycloalkenyl, C₅₋₇cycloalkenyl C₁₋₁₀ alkyl, halogen, (CR₁₀R₂₀)_(n)OR₆, (CR₁₀R₂₀)_(n)SH,(CR₁₀R₂₀)_(n)S(O)_(m)R₇, (CR₁₀R₂₀)_(n)NHS(O)₂R₇, (CR₁₀R₂₀)_(n)NR₄R₁₄,(CR₁₀R₂₀)_(n)CN, (CR₁₀R₂₀)_(n)S(O)₂NR₄R₁₄, (CR₁₀R₂₀)_(n)C(Z)R₆,(CR₁₀R₂₀)_(n)OC(Z)R₆, (CR₁₀R₂₀)_(n)C(Z)OR₆, (CR₁₀R₂₀)_(n)C(Z)NR₄R₁₄,(CR₁₀R₂₀)_(n)NR₁₀C(Z)R₆, (CR₁₀R₂₀)_(n)NR₁₀OC(═NR₁₀)NR₄R₁₄,(CR₁₀R₂₀)_(n)OC(Z)NR₄R₁₄, (CR₁₀R₂₀)_(n)NR₁₀C(Z)NR₄R₁₄, or(CR₁₀R₂₀)_(n)NR₁₀C(Z)OR₇; R₃ is a C₁₋₄ alkyl, halo-substituted C₁₋₄alkyl, C₁₋₄ alkenyl, C₁₋₄ alkynyl, C₃₋₇ cycloalkyl, C₅₋₇ cycloalkenyl,aryl, arylC₁₋₄ alkyl, heteroaryl, heteroarylC₁₋₄ alkyl, heterocyclyl,heterocyclylC₁₋₄ alkyl, (CR₁₀R₂₀)_(v)OR₇, (CR₁₀R₂₀)_(v)S(O)_(m)R₇,(CR₁₀R₂₀)_(v)NHS(O)₂R₇, or (CR₁₀R₂₀)_(v)NR₄R₁₄; and wherein the aryl,arylalkyl, heteroaryl, or heteroaryl alkyl may be optionallysubstituted; R₄ and R₁₄ is each independently selected from hydrogen oroptionally substituted C₁₋₄ alkyl, optionally substituted aryl oroptionally substituted aryl-C₁₋₄ alkyl, or together with the nitrogenwhich they are attached form a heterocyclic ring of 5 to 7 members whichring optionally contains an additional heteroatom selected from oxygen,sulfur or NR₉; R₅ is hydrogen, C₁₋₄ alkyl, C₂₋₄ alkenyl, C₂₋₄ alkynyl orNR₄R₁₄, excluding the moieties SR₅ being SNR₄R₁₄, S(O)₂R₅ being SO₂H andS(O)R₅ being SOH; R₆ is hydrogen, C₁₋₁₀ alkyl, C₃₋₇ cycloalkyl,heterocyclyl, heterocyclyl C₁₋₁₀ alkyl, aryl, arylC₁₋₁₀ alkyl,heteroaryl or heteroarylC₁₋₁₀ alkyl, wherein these moieties may beoptionally substituted; R₇ is a C₁₋₆alkyl, aryl, arylC₁₋₆alkyl,heterocyclic, heterocyclylC₁₋₆ alkyl, heteroaryl, or heteroarylC₁₋₆alkylmoiety; and wherein each of these moieties may be optionallysubstituted; R₈ is hydrogen, C₁₋₄ alkyl, halo-substituted C₁₋₄ alkyl,C₂₋₄ alkenyl, C₂₋₄ alkynyl, C₃₋₇ cycloalkyl, C₅₋₇ cycloalkenyl, aryl,arylC₁₋₄ alkyl, heteroaryl, heteroarylC₁₋₄ alkyl, heterocyclyl,heterocyclylC₁₋₄ alkyl, (CR₁₀R₂₀)_(t)OR₇, (CR₁₀R₂₀)_(t)S(O)_(m)R₇,(CR₁₀R₂₀)_(t)NHS(O)₂R₇, or (CR₁₀R₂₀)_(t)NR₄R₁₄; and wherein the aryl,arylalkyl, heteroaryl, heteroaryl alkyl may be optionally substituted;R₉ is hydrogen, C(Z)R₆ or optionally substituted C₁₋₁₀ alkyl, optionallysubstituted aryl or optionally substituted aryl-C₁₋₄ alkyl; R₁₀ and R₂₀are each independently selected from hydrogen or C₁₋₄ alkyl; R₁₁ is aC₁₋₄ alkyl, halo-substituted C₁₋₄ alkyl, C₂₋₄ alkenyl, C₂₋₄ alkynyl,C₃₋₇ cycloalkyl, C₅₋₇ cycloalkenyl, aryl, arylC₁₋₄ alkyl, heteroaryl,heteroarylC₁₋₄ alkyl, heterocyclyl, heterocyclylC₁₋₄ alkyl,(CR₁₀R₂₀)_(t)OR₇, (CR₁₀R₂₀)_(t)S(O)_(m)R₇, (CR₁₀R₂₀)_(t)NHS(O)₂R₇, or(CR₁₀R₂₀)_(v)NR₄R₁₄; and wherein the aryl, arylalkyl, heteroaryl, orheteroaryl alkyl may be optionally substituted; Z is oxygen or sulfur; nis 0, or an integer having a value of 1 to 10; m is 0, or the integer 1or 2; v is 0, or an integer having a value of 1 or 2; t is an integerhaving a value of 1 to 3; or a pharmaceutically acceptable salt thereof.2. The compound according to claim 1 wherein R₁ is an optionallysubstituted phenyl or naphthyl.
 3. The compound according to claim 2wherein the substituent is independently selected from halogen, alkyl,hydroxy, alkoxy, amino, or halosubstituted alkyl.
 4. The compoundaccording to claim 3 wherein the substituent is a halogen.
 5. Thecompound according to claim 1 wherein R₂ is an optionally substituted,arylalkyl, C₃₋₇ cycloalkyl, heteroarylC₁₋₁₀ alkyl, or heterocyclylC₁₋₁₀alkyl.
 6. The compound according to claim 5 wherein R₂ is an optionallysubstituted, benzyl, phenethyl, cyclohexyl, pyrrolindylmethyl,morpholino ethyl, morpholinopropyl, C₁₋₄ alkyl, or pyridylethyl.
 7. Thecompound according to claim 5 wherein the substitutent is independentlyhalogen, alkyl, hydroxy, alkoxy, amino, or halosubstituted alkyl.
 8. Thecompound according to claim 6 wherein the substituent is a halogen, oralkyl.
 9. The compound according to claim 1 which is:1-Cyclohexyl-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one;1-Isopropyl-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one;(+/−)-1-[1-(Phenyl)ethyl]-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one;(R)-1-[1-(Phenyl)ethyl]-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one;(S)-1-[1-(Phenyl)ethyl]-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one;1-(2-Phenethyl)-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one;1-Benzyl-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one;1-(2-morpholin-4-yl-ethyl-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one;1-[2-(2-pyridinyl)ethyl]-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one;trifluoroacetate1-(2-morpholin-4-yl-propyl)-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one;trifluoroacetate1-(1-methylpyrrol-2-ylethyl)-5-phenyl-3,4-dihydro-(1H)-quinazolin-2-one;or a pharmaceutically acceptable salt thereof.
 10. A pharmaceuticalcomposition comprising an effective amount of a compound according toclaim 1 and a pharmaceutically acceptable carrier or diluent.
 11. Amethod of treating a CSBP/RK/p38 kinase mediated disease in a mammal inneed thereof, which method comprises administering to said mammal aneffective amount of a compound of Formula (I) according to claim
 1. 12.The method according to claim 11 wherein the CSBP/RK/p38 kinase mediateddisease is psoriatic arthritis, Reiter's syndrome, gout, traumaticarthritis, rubella arthritis and acute synovitis, rheumatoid arthritis,rheumatoid spondylitis, osteoarthritis, gouty arthritis, sepsis, septicshock, endotoxic shock, gram negative sepsis, toxic shock syndrome,cerebral malaria, meningitis, ischemic and hemorrhagic stroke,neurotrauma/open or closed head injury, asthma, adult respiratorydistress syndrome, chronic pulmonary inflammatory disease, chronicobstructive pulmonary disease, silicosis, pulmonary sarcososis, boneresorption disease, osteoporosis, restenosis, cardiac, brain and renalreperfusion injury, thrombosis, glomerularnephritis, chronic renalfailure, diabetes, diabetic retinopathy, macular degeneration, graft vs.host reaction, allograft rejection, inflammatory bowel disease, Crohn'sdisease, ulcerative colitis, neurodegenrative disease, muscledegeneration, diabetic retinopathy, macular degeneration, tumor growthand metastasis, angiogenic disease, eczema, contact dermatitis,psoriasis, sunburn, or conjunctivitis.
 13. A method of treating thecommon cold or respiratory viral infection caused by human rhinovirus(HRV), other enteroviruses, coronavirus, influenza virus, parainfluenzavirus, respiratory syncytial virus, or adenovirus in a human in needthereof which method comprises administering to said human an effectiveamount of a compound of Formula (I) according to claim
 1. 14. The methodaccording to claim 13 wherein the respiratory viral infectionexacerbates asthma, chronic bronchitis, chronic obstructive pulmonarydisease, otitis media, or sinusitis.
 15. A method of treatment of smokeinduced airway inflammation in a human in need thereof, which comprisesadministering to said human an effective amount of a compound accordingto claim
 1. 16. The method according to claim 15 wherein the smokeinduced airway inflammation is caused by inhalation of cigarette smoke,inhalation of smoke produced from a burning plant material, orinhalation of burning smoke from fossil fuels.
 17. The method accordingto claim 15 wherein the smoke induced airway inflammation exacerbates apre-existing asthmatic condition, a pre-existing chronic bronchitis, orpre-existing chronic obstructive pulmonary disease in said human.
 18. Amethod of treatment, for inflammation enhanced cough in a mammal in needthereof, which comprises administering to said mammal an effectiveamount of a compound according to claim
 1. 19. The method according toclaim 18 wherein the inflammation enhanced cough is cough variantasthma, or eosinophilic bronchitis.
 20. The method according to claim 13wherein the compound of Formula (I) is administered with a secondtherapeutic agent.
 21. The method according to claim 20 wherein thesecond therapeutic agent is an anti-tussive; an antihistamine; asteroid; a PDE₄ agent, an antibiotic; an anti-inflammatory agentselected from an NSAID, a COX-1 or COX-2 inhibitor, ASA, orindomethacin; an antiviral agent ribavirin, amantidine, rimantidine,Pleconaril, AG 7088, or BTA-188; a decongestant; an influenzaneuraminidase inhibitor selected from zamanivar (Relenza), oseltamivir(Tamiflu) or RWJ-270201.
 22. The method according to any one of claims13 to 19 wherein the compound of Formula (I) is administered orally,topically (intranasal) or via inhalation (aerosol), or both topicallyand via inhalation.
 23. The method according to claim 15 wherein thecompound of Formula (I) is administered with a second therapeutic agent.24. The method according to claim 23 wherein the second therapeuticagent is an antitussive; an antihistamine; a steroid; a PDE₄ agent, anantibiotic; an anti-inflammatory agent selected from an NSAID, a COX-1or COX-2 inhibitor, ASA, or indomethacin; an antiviral agent ribavirin,amantidine, rimantidine, Pleconaril, AG 7088, or BTA-188; adecongestant; an influenza neuraminidase inhibitor selected fromzamanivar (Relenza), oseltamivir (Tamiflu) or RWJ-270201.
 25. The methodaccording to any claim 15 wherein the compound of Formula (I) agent isadministered orally, topically (intranasal) or via inhalation (aerosol),or both topically and via inhalation.
 26. The method according to claim18 wherein the compound of Formula (I) is administered with a secondtherapeutic agent.
 27. The method according to claim 26 wherein thesecond therapeutic agent is an antitussive; an antihistamine; a steroid;a PDE₄ agent, an antibiotic; an anti-inflammatory agent selected from anNSAID, a COX-1 or COX-2 inhibitor, ASA, or indomethacin; an antiviralagent ribavirin, amantidine, rimantidine, Pleconaril, AG 7088, orBTA-188; a decongestant; an influenza neuraminidase inhibitor selectedfrom zamanivar (Relenza), oseltamivir (Tamiflu) or RWJ-270201.
 28. Themethod according to any claim 18 wherein the compound of Formula (I)agent is administered orally, topically (intranasal) or via inhalation(aerosol), or both topically and via inhalation.